HK1235693A - Methods and compositions to treat hemorrhagic conditions of the brain - Google Patents

Description

Methods and compositions for treating hemorrhagic conditions of the brain

The present application is a divisional application of the following applications: application date: 2011, 2 month, 22 days; application No.: 201180018383. X; the invention name is as follows: as above.

Cross Reference to Related Applications

This application claims the benefit of priority from U.S. provisional application No.61/306,758 filed on 22/2/2010, which is incorporated herein by reference in its entirety.

Technical Field

The invention relates to methods and compositions for treating hemorrhagic conditions of the brain.

Background

The central nervous system is a bilateral and substantially symmetrical structure with the following 7 parts: spinal cord, medulla oblongata, pons, cerebellum, midbrain, diencephalon, and hemispheres. FIG. 1 shows a side view of a human brain from Stedman's Medical Dictionary, 27 th edition, plate 7at A7 (2000).

The spinal cord, the most caudal portion of the central nervous system, receives and processes sensory information from the skin, joints and muscles of the limbs and trunk and controls the movement of the limbs and trunk. It is subdivided into the cervical, thoracic, lumbar and sacral regions. The spinal cord continues forward as a brainstem consisting of medulla, pons and the middle brain. The brainstem receives sensory information from the skin and muscles of the head and provides motion control for the head muscles. It also transmits information from the spinal cord to the brain and from the brain to the spinal cord, and regulates the level of wakefulness and wakefulness through the reticular structure. The brain stem contains several cell body aggregates (cranial nerve nuclei). Some of these sets receive information from the skin and muscles of the head; others control the movement output of facial, neck and eye muscles. Other sets are dedicated to being from special senses: auditory, balance, and gustatory information. (Kandel, E. et al, Principles of neurol science, 4 th edition, page 8, 2000).

Medulla oblongata, located directly in the front of the spinal cord, includes several centers responsible for important autonomic functions (such as digestion, respiration, and heart rate controls) (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

The pons, located in the anterior part of the medulla, transmit information about movement from the hemispheres of the brain to the cerebellum (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

The cerebellum is located behind the pons and is connected to the brainstem by several main fiber bundles called feet. The cerebellum regulates the strength and range of motion and participates in the learning of motor skills. It also contributes to learning and cognition (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

The midbrain, located in the front of the pons, controls many sensory and motor functions, including coordination of eye movement and visual and auditory reflexes (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

The diencephalon is located in the anterior part of the midbrain and contains two structures. One structure, the thalamus, processes most of the information that reaches the cerebral cortex from the rest of the central nervous system and participates in other functions, including motor control, autonomic function, and cognition. Another structure, the hypothalamus, regulates autonomic, endocrine, and visceral functions (Kandel, e. et al, Principles of neural Science, 4 th edition, page 8, 2000).

The hemispheres of the brain consist of a highly convoluted outer layer (the cerebral cortex) and three deeply buried structures-the basal ganglia involved in the regulation of motor performance; hippocampus which participates in learning and memory storage; and amygdala-component that coordinates autonomic and endocrine responses to emotional states (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

The cerebral cortex is divided into 4 lobes: frontal, parietal, temporal and occipital lobes. The surface of the hemisphere contains many concave lines or grooves, called fissures and furrows. The portion of the brain located between these fovea lines is called the gyrus or gyri. A lateral cerebral fissure (sierviels fissure) separates the temporal lobe from the frontal lobe. The central groove (Rolandic sulcus) separates the frontal and parietal lobes (Kandel, E. et al, Principles of Neural Science, 4 th edition, page 8, 2000).

1. Meninges of brain

The meninges (three distinct connective tissue membranes that surround and protect the brain and spinal cord) are named (from the outer to the inner layer) dura mater, arachnoid mater, and pia mater. Fig. 2 shows a diagrammatic sagittal plane of the human brain (j.g. mouse, relative neuroanatomi & Functional Neurology, 18 th edition, page 46, 1982).

1.1. Dura mater

The dura mater is a dense fibrous structure that covers the brain and spinal cord. It has an inner meningeal layer and an outer periosteal layer or endosteal layer. The dura mater on the brain is generally confluent, except where they separate to provide a venous sinus compartment and where the inner layer forms a space between the brain portions. The outer layer adheres firmly to the inner surface of the skull and sends the vascular and fibrous extensions into the bone itself. Around the edge of the occipital foramen (the large opening of the channel from the cranial cavity to the spinal cavity in the base of the skull), it adheres tightly to the bone and is uninterrupted by the dura mater.

The dura mater consists of fibroblasts, abundant extracellular collagen and a small amount of elastic fibers arranged in a flat layer, which is incompletely divided into two layers closely connected together by lacunar space and blood vessels: the inner layer (meningeal layer) and the outer layer (endosteal layer), with some exceptions, are separated to form the sinuses for venous blood passage or to form the space between the brain portions. The outer surface of the dura is rough and fibrillated (consisting of fibers) and adheres closely to the inner surface of the bone, with adhesion most evident opposite the cranial sutures (the immobile joints between the bones of the cranium or cranium). The endosteal layer is the inner periosteum of the skull and contains the blood vessels that supply them. The meningeal layer is lined on its inner surface with a layer of unique elongated flattened fibroblasts, which have been called dural border layer cells. Collagen is not present in this layer and cells are not connected by cell junctions. They are often separated by extracellular spaces filled with amorphous, non-filamentous material. The meningeal layer further comprises two layers: a dense layer and a loose layer; the former generally contains strong fibrous tissue and a few blood vessels, but the latter contains some blood vessels. Figure 3 is a cross-sectional view of the 3 meningeal layers covering the brain (Haines, D.E., an atomic record 230:3-21,1991). The dura mater sends four projections inward that divide the cranial cavity into a series of freely communicating compartments and provide further protection to different parts of the brain.

The dural processes that protrude into the cranial cavity are formed by the re-repetition of the inner (or meningeal) layer of the membrane. These protrusions include: (1) the brain sickle, (2) the cerebellum, (3) the cerebellum sickle and (4) the diaphragm.

The sickle is a powerful, arcuate projection in the form of a sickle that descends vertically at the longitudinal cleft between the hemispheres of the brain. It narrows anteriorly where it joins the ethmoid (bone at the base of the cranium and at the nasal root) at the comb (triangular midline prominence of the ethmoid); and broad at the back where it joins the upper surface of the cerebellum (the curved fold of dura covering the upper surface of the cerebellum). Its upper edge is convex and connects with the inner surface of the skull at the midline, as far back as possible as the inion protuberance; it contains the superior sagittal sinus. Its inferior border is free and concave and contains the inferior sagittal sinus.

The cerebellum is an arc-shaped layer that is elevated in the middle and slopes downward toward the periphery. It covers the upper surface of the cerebellum and supports the occipital lobes of the brain. Its leading edge is free and concave and incorporates a large oval opening (the curtal notch) for the passage of the brain foot (the large cortical nerve fiber bundle passing longitudinally on each side of the midline in the ventral aspect of the midbrain) as well as descending sensory and autonomic fibers and other fiber bundles. The cerebellum is attached posteriorly by its convex edge to the transverse ridge on the inner surface of the occiput and there surrounds the transverse sinus; and anteriorly attached to the superior corner of the temporal bony rock on either side, surrounding the supralithologic sinus. At the vertex of the temporal bony rock portion, the free and attached margins coincide and intersect each other, continuing anteriorly to be fixed to the anterior and posterior bedprocesses, respectively. The posterior edge of the sickle is attached to the midline of its upper surface. The direct sinus is located at the junction of the sickle of the brain and the cerebellum.

The cerebellar sickle is a small triangular protrusion of the dura mater separating two cerebellar hemispheres. The base of which is attached above to the lower and back parts of the screen; and the lower part of the vertical ridge whose trailing edge is attached to the inner surface of the occiput splits. As it descends, the sickle sometimes splits into two smaller folds which disappear on the sides of the large pillow opening.

The saddletree is a small circular horizontal fold that covers and almost completely covers the pituitary gland (pituitary gland) in the sphenoid saddles (saddle-like protuberances on the upper surface of the sphenoid bone of the skull, lying within and bisecting the fovea cranialis); the central opening of variable size penetrates the funnel (the funnel-shaped extension of the hypothalamus that connects the pituitary to the base of the brain).

The arteries of the dura are numerous. The meningeal branch of the anterior and posterior ethmoid arteries and the internal carotid artery and the branch from the artery in the meninges supply the dura mater of the anterior cranial fossa. The meningeal middle and accessory arteries of the internal jaw artery; a branch from the ascending pharyngeal artery into the skull through the ruptured aperture; branches from the internal carotid artery and the return branches from the lacrimal artery supply the dura mater of the cranial fossa. Meningeal branches from the occipital artery, one entering the skull via the jugular vein foramen and the other entering the skull via the mastoid foramen; posterior meningeal artery from vertebral artery; an adventitious branch of the meninges from the ascending pharyngeal artery into the skull via the jugular vein foramen and the sublingual nerve canal; and the dura mater, which branches from the artery in the meninges supply the posterior cranial fossa.

The veins returning blood from the dura mater are anastomosed to the plateaus veins or terminate in multiple sinuses. Many meningeal veins do not open directly into the sinuses, but open indirectly through a series of ampullas (called venous fossae). These ampules are present on either side of the superior sagittal sinus, particularly in the central portion thereof, and are often invaginated by arachnoid particles; they are also present in the vicinity of the transverse and direct sinuses. They communicate with the underlying cerebral veins and also with the parapet vein and the vena cava.

The dural nerve is a fibril (filamentt) derived from the trigeminal ganglion, glossopharyngeal ganglion, vagus nerve ganglion, second and third spinal ganglia, sphenopalatine ganglion, ear ganglion, and supracervical ganglion and provides unmyelinated and myelinated sensory and autonomic fibers.

1.2. Arachnoid membrane

The middle cerebral lamina (arachnoid membrane) is a delicate avascular membrane located between the pia and dura mater. It is separated from the superior dura mater by the subdural space and from the inferior pia mater by the subarachnoid space containing cerebrospinal fluid.

The arachnoid membrane consists of an outer cell layer of low cubic mesothelium (low cubic mesothelium). There is a space of variable thickness which is filled with cerebrospinal fluid and is traversed by trabeculae and membranes composed of collagen fibrils and cells like fibroblasts. The lining and trabeculae are covered with more or less low-type cubic mesothelium that is flattened into a paving stone type (pavement type) at multiple places and mixed with the cells of the pia mater on the deep lining. The arachnoid membrane also contains plexuses in the motor root that originate from the trigeminal, facial and cranial accessory nerves.

The cranial portion of the arachnoid (the brain arachnoid) falls loosely into the brain and does not sag into the sulcus (depressions or fissures on the surface of the brain) between the gyrus (raised folds or plateaus on the surface of the brain) nor into fissures, except for a longitudinal fissure and several other larger sulci and fissures. On the surface of the brain, the arachnoid is thin and transparent; at the base, it is thicker. It is slightly opaque towards the central part of the brain, where it extends between the two temporal lobes in front of the pons, leaving a large space between the pons and the brain.

The arachnoid surrounds the cranial and spinal nerves and encases them in a loose sheath until their tips exit the skull.

Arachnoid membrane

The subarachnoid space (subarachnoid cavity) or subarachnoid space (subarachnoid space), which is a space between the arachnoid extracellular layer and the pia mater, is occupied by a tissue composed of a fragile connective tissue trabecula and an interconnecting channel containing cerebrospinal fluid therein. This cavity is small on the surface of the hemisphere; on the tip of each gyrus, the pia mater and the arachnoid are in close contact, but the triangular spaces are left in the sulcus between the gyrus where there is subarachnoid trabecular tissue, as the pia mater sinks into the sulcus, while the arachnoid bridges these triangular spaces between the gyrus. In certain parts of the base of the brain, the arachnoid is separated from the pia mater by wide gaps that are in free communication with each other and are designated as subarachnoid pools; subarachnoid tissue in these pools is less abundant.

Subarachnoid pool (cisternae subarachnoid)

The cisterna magna (cisterna magna) is triangular in the sagittal section and results from bridging the arachnoid space across these gaps between the medulla oblongata and the surface of the cerebellar hemisphere; it is connected with subarachnoid space of spinal cord at the level of large hole of pillow.

The pontine is a large cavity on the ventral surface of the brain bridge. It contains the basilar artery and is joined to the subarachnoid space of the spinal cord behind the pons and to the cisterna magna; in front of the pons, it is connected to the interpeduncular pool.

The interpeduncular cistern (basal cistern) is a broad cavity in which the arachnoid extends across both temporal lobes. It surrounds the brain feet and structures contained in the interpeduncular fossa, and contains the cerebral arterial loop. Anteriorly, the interpeduncular cistern extends anteriorly across the visual cross, forming a pool of crossings, and onto the upper surface of the corpus callosum. The arachnoid stretch across one hemisphere to the other just below the free edge of the sickle and thus leaves a cavity in which the anterior cerebral artery is located. The lateral fossa cerebri is formed in front of either temporal lobe by a spider web bridging across the lateral fissure. This cavity contains the middle cerebral artery. The cisterna magna occupies the space between the callosomal pressor portion and the upper surface of the cerebellum; it extends between the layers of choroidal tissue in the third ventricle and contains the great cerebral veins.

The subarachnoid space is in communication with the general ventricular cavity of the brain through 3 openings; one opening (massadehole) is in the midline at the inferior portion of the roof of the fourth ventricle; the other two (luschka holes) are at the lateral crypt end of this ventricle, behind the upper root of the glossopharyngeal nerve.

The arachnoid villi is a clustered extension of the pia mater, which protrudes through the meningeal layer of the dura mater and has a thin limiting membrane. The clustered extensions of the pia mater, consisting of numerous arachnoid villi that penetrate the dural venous sinus and cause the transfer of cerebrospinal fluid into the venous system, are called arachnoid granules.

The arachnoid villi represents invasion of the dura by the arachnoid, and therefore, the arachnoid mesothelial cells are present just below the vascular endothelium of the great dural sinus. Each pile consists of the following parts: (1) inside is a core of subarachnoid tissue, which is joined to the reticulum of total subarachnoid tissue via the narrow pedicle through which the villi attach to the arachnoid; (2) surrounding this tissue is the arachnoid layer that defines and surrounds the subarachnoid tissue; (3) in this arachnoid layer is a thinned fossa wall, which is separated from the arachnoid by a potential cavity corresponding to and joining the potential subdural space; and (4) if the villus protrudes into the sagittal sinus, it will be covered by a significantly thinner sinus wall, which may consist of the endothelium only. Fluid injected into the subarachnoid space will enter these villi. This fluid passes from the villi into the antrum venosus into which the villi protrude.

1.3. Soft film

The pia mater is a thin connective tissue membrane applied to the surface of the brain and spinal cord. Blood vessels supplying the brain enter the brain through the pia. The pia mater is absent at the maxdie and two luschka holes and is crossed by all blood vessels as they enter or leave the nervous system and is therefore considered to be an incomplete membrane. In the perivascular space, the mesothelial lining on the surface of the pia mater, which serves as the outer surface of the compartment, enters; at variable distances from the outside, these cells become unrecognizable and apparently absent, being replaced by glial units. The inner wall of the perivascular space generally appears to be covered a distance by mesothelial cells, reflected as the vessels from the arachnoid coating of these vascular channels as they traverse the subarachnoid space.

The pia mater (pia mater encephali), the pia of the brain, covers the entire surface of the brain, sags between the gyrus and cerebellar layers, and intussuscepts to form the choroidal tissue of the third ventricle and choroid plexuses of the lateral and third ventricles. When the pia mater passes over the parietal aspect of the fourth ventricle, it forms the choroidal tissues and choroid plexus of the fourth ventricle. On the cerebellum, the membrane is more fragile; the blood vessels from their deep surface are short and their relationship to the cortex is not so intimate.

The pia mater forms the sheath of the cranial nerve.

2. Circulation of the brain

The willis ring at the base of the brain is the main arterial anastomotic trunk of the brain. Blood reaches it mainly via the spinal and internal carotid arteries (see fig. 4); anastomosed between branches of the Willis's loop on the hemisphere and occurs via extracranial arteries which penetrate the skull via a plurality of orifices.

The circle of Willis is formed by the anastomosis between the internal carotid artery, the basilar artery, the anterior cerebral artery, the anterior communicating artery, the posterior cerebral artery and the posterior communicating artery. The internal carotid artery terminates in the anterior cerebral artery and the middle cerebral artery. Near its end, the internal carotid artery creates a posterior communicating artery, which caudally joins the posterior cerebral artery. The anterior cerebral artery is connected via the anterior communicating artery.

Blood supply to the cerebral cortex is mainly by means of the cortical branches of the anterior, middle and posterior cerebral arteries, which reach the cortex in the pia mater. FIG. 5 shows a schematic diagram of the arterial supply to the cerebral cortex, where 1 is the orbitofrontal artery; 2 is the anterior Rolando artery; 3 is a Rolando artery; 4 is the parietal anterior artery; 5 is the parietal posterior artery; 6 is the inner canthus artery; 7 is the posterior temporal artery; 8 is the anterior temporal artery; 9 is the orbital artery; 10 is the frontal polar artery; 11 is an artery at the margin of the corpus callosum; 12 is the internal posterior artery; and 13 is a peripheral artery of the corpus callosum. (relative neuroanatomi & Functional Neurology, 18 th edition, page 50, 1982).

The lateral surface of each hemisphere is supplied primarily by the middle cerebral artery. The medial and diaphragmatic supply of the hemispheres is supplied primarily by the anterior and posterior cerebral arteries.

The middle cerebral artery (the terminal branch of the internal carotid artery) enters the cerebral lateral fissure and splits into cortical branches adjacent to the frontal, temporal, parietal and occipital lobes. The penetrating arteriole (the lenticulate artery) originates from the base of the middle cerebral artery to supply the inner sac structure and adjacent structures.

The anterior cerebral artery extends medially from its origin from the internal carotid artery into the paranemia to the corpus callosum knee, where it switches next to the corpus callosum. It creates branches that reach these lobes along the medial side of the medial frontal and parietal lobes and reach the adjacent cortex.

The posterior cerebral artery originates at its anterior end from the basilar artery, usually at the midbrain level, curves around the dorsum of the brain, and branches to the medial side of the temporal lobe and diaphragm and to the medial occipital lobe. The branches include the brachial artery and the fenestrated branches to the posterior and hypothalamus.

The basilar artery is formed by the junction of the vertebral arteries. It is supplied to the upper brainstem through the short, short and long branches of the medial collateral channel.

The midbrain is supplied by the basilar, posterior cerebral and superior cerebellar arteries. The pons are supplied by the basilar, anterior, inferior and superior cerebellar arteries. The medulla oblongata is supplied by the vertebral, anterior, posterior, inferior and posterior cerebellar arteries and the basilar artery. The cerebellum is supplied by the cerebellar arteries (the upper, anterior and lower cerebellar arteries and the posterior and lower cerebellar arteries).

The choroid plexus of the third and lateral ventricles is supplied by branches of the internal carotid and posterior cerebral arteries. The choroid plexus of the fourth ventricle is supplied by the posterior lower cerebellum artery.

Venous outflow from the brain is primarily to the epidural sinus (a vascular access located inside the dural solid). The epidural sinus does not contain a valve and is, for the most part, triangular in shape. The superior sagittal sinus is located in the sickle of the brain.

3. Hemorrhagic conditions of the brain

Annually, over 2,000,000 people worldwide suffer spontaneous or traumatic intracerebral hemorrhage ("ICH"), both of which have poor drug outcomes, are difficult to treat, and have high mortality and morbidity. Spontaneous ICH has the highest morbidity and mortality of all strokes. Chronic subdural hematoma ("SDH") is also a common neurosurgical problem. There is little epidemiological data on its incidence, but neurosurgical practice has shown that the incidence is more than 30 per 100,000 people per year.

Treatment of hemorrhagic encephalopathies, including removal by craniotomy or less invasive surgery by drilling, are all of variable effectiveness. One important complication of these forms of bleeding is postoperative rebleeding, which occurs in 10-30% of cases and increases morbidity and mortality. Postoperative bleeding can also occur following intracranial surgery for other conditions such as brain tumors, epilepsy, infections, and cerebrovascular malformations.

3.1. Intracerebral hemorrhage (ICH)

The term "non-traumatic ICH" as used herein refers to bleeding into the brain parenchyma caused by or associated with trauma and particularly traumatic injuries, which may extend into the cerebral ventricles and, in some cases, into the subarachnoid space. The term "traumatic ICH" as used herein refers to such bleeding caused by or associated with trauma and especially traumatic injuries.

Idiopathic and traumatic ICH are important causes of morbidity and mortality worldwide. The estimated annual incidence of spontaneous ICH is 15-30/100,000 in the population. Treatment consisted of careful care support, treatment of increased intracranial pressure, and surgical clearance of hematomas in selected cases. These treatments are not very effective; mortality rates exceed 50% and survivors often have severe morbidity. One cause of poor drug outcome is bleeding before or after surgical removal of the hematoma or after surgical removal of the tumor, infection, or vascular malformation. Hematoma growth occurred in up to 70% of patients imaged within three hours of ICH. In addition, bleeding spread is an independent determinant of death and disability. In addition to ICH growth, other predictors of poor drug outcome include age, baseline volume of bleeding, glasgow coma scale score, intraventricular hemorrhage and subterminal location. Postoperative rebleeding also occurs in up to 6% of patients, but is more common in early surgery (40% within 4 hours of shock). However, patients who undergo surgery at an early stage may benefit most from surgery, and thus reducing the risk of early rebleeding may be critical. There are reports in the medical literature of the administration of drugs to prevent bleeding in patients who have undergone surgery to remove ICH. Systemic (intravenous or generally meaning the body) administration of an antifibrinolytic agent or activated factor VIIa to reduce rebleeding does not improve the dosing outcome, partly due to systemic side effects of the drug. For example, ICH patients treated with recombinant factor VII present an increased risk of arterial thromboembolic complications (most common cerebral infarction and myocardial ischemia) as indicated by elevated troponin I concentrations (Diringer, MN et al, Stroke 13: 850-.

Traumatic ICH caused by Traumatic Brain Injury (TBI) is even more common than spontaneous ICH. Approximately 10% of TBI (1,400,000 annual us cases) is associated with ICH requiring surgery. The literature does not adequately record how often recurrent bleeding occurs in the contused brain following trauma. According to some estimates, post-traumatic rebleeding occurs in up to 10% of patients and is therefore a serious concern for neurosurgeons.

Pathogenesis of ICH

ICH commonly occurs in the basal ganglia, thalamus, brainstem (predominantly pons), hemispheres and cerebellum. The expansion into the ventricles of the brain is accompanied by a profound hematoma. Edema parenchyma, often discolored by degradation products of hemoglobin, are seen adjacent to the blood clot. Histological sections were characterized by the presence of edema, neuronal damage, macrophages and neutrophils within the area surrounding the hematoma. The bleeding spreads between the white matter cutting planes, causing some disruption of the brain structure, and leaves multiple turns of intact neural tissue inside and around the hematoma. This pattern of expansion explains the presence of viable and recoverable neural tissue in close proximity to the hematoma.

Intraparenchymal hemorrhage is commonly caused by rupture of small penetrating arterioles originating from the basilar artery or from the anterior, middle or posterior cerebral artery. Degenerative changes in the arteriolar wall caused by chronic hypertension reduce compliance, weaken the arteriolar wall, and increase the likelihood of spontaneous rupture. Studies have shown that most bleeding occurs at or near the bifurcation of the affected artery, where significant degeneration of the media and smooth muscle can be seen.

The ICH extends over time. Studies using Computed Tomography (CT) scanning have shown that hematomas spread in 26% of patients within 1 hour after the initial CT scan and in another 12% of patients within 20 hours. Other studies have shown that hematomas spread among 20% of ICH patients, occur in 36% of patients present within 3 hours after bleeding onset and in 11% of patients present more than 3 hours after onset. This expansion has been attributed to continued bleeding from the original source and to mechanical damage to the surrounding blood vessels. Acute hypertension, local coagulation deficiencies, or both may accompany the expansion of hematomas.

The presence of hematoma initiates edema and neuronal damage within the surrounding parenchyma. Fluid begins to immediately accumulate in the area around the hematoma, edema usually increases up to 5 days, and edema has been observed for up to 2 weeks after stroke. Early edema surrounding the hematoma results from the release and accumulation of osmotically active serum proteins from the blood clot. Angiogenic edema and cytotoxic edema follow, due to blood brain barrier disruption, sodium pump failure, and neuronal death.

The delay in blood brain barrier destruction and the formation of cerebral edema following ICH suggests the possible presence of secondary mediators of nerve damage and edema. Blood and plasma products are generally thought to mediate most of the secondary processes initiated after ICH. It is uncertain whether ischemia occurs due to mechanical compression or some chemical action of the bleeding in the area surrounding the hematoma. Neuronal death in the peri-hematoma region is predominantly necrotic (some studies indicate that there is also programmed cell death (apoptosis)).

Classification of ICH

ICH is classified as primary or secondary depending on the underlying cause of the bleeding. Primary ICH, accounting for 78-88% of cases, is derived from spontaneous rupture of small blood vessels damaged by chronic hypertension, amyloid angiopathy or some other cause. Secondary ICH occurs in a few patients with vascular abnormalities (such as arteriovenous malformations and aneurysms), tumors, or impaired clotting.

Biomarkers for ICH

The surface of all cells in the body is coated with specialized protein receptors that have the ability to selectively bind to or adhere to other signaling molecules (Weiss and Littman, Cell,76:263-74, 1994). These receptors and the molecules to which they bind are used to communicate with other cells and to perform the correct cellular functions within the body. Each cell type has a specific receptor combination or label on the surface that distinguishes them from other cell types.

The term "biomarker" (or "biomarker") as used herein refers to a peptide, protein, nucleic acid, antibody, gene, metabolite, or any other substance that serves as an indicator of a biological state. It is a characteristic of cellular or molecular indicators that are objectively measured and evaluated as pharmacological responses to normal biological processes, pathogenic processes, or therapeutic interventions. The term "indicator" as used herein refers to any substance, number or ratio that can reveal a relative change as a function of time, resulting from a series of observed facts; or a signal, sign, marker, sign or symptom that is visible or evidences its presence or presence. Once the proposed biomarker has been validated, it can be used to diagnose disease risk, the presence of disease in an individual, or to tailor disease therapy (selection of drug treatment or administration regimen) in an individual. Biomarkers can be used as surrogates for natural endpoints (e.g., survival or irreversible morbidity) in evaluating potential drug therapies. If a treatment alters such a biomarker and the alteration is directly associated with improved health, such a biomarker may serve as a surrogate to assess clinical benefit. Clinical endpoints are variables that can be used to measure how well a patient feels, functions, or survives. Surrogate endpoints may be biomarkers intended to replace clinical endpoints; these biomarkers are validated to predict clinical endpoints with acceptable confidence levels for regulatory agencies and clinical circles.

Studies have shown that ICH affects a coordinated network of gene expression within the inflammatory system, anti-inflammatory system and neuronal signaling system in human hematoma surrounding tissue within the first day of bleeding. The hematoma surrounding tissue includes gray matter structures and white matter structures. Edema surrounding a hematoma with significant impact is an almost universal complication of ICH. Molecular networks of proinflammatory signaling begin with the cytokines interleukin 1 beta (IL-1 beta), IL-8, IL-6 receptor, CCR1, CXCL2/MIP2, and CXCL 3. These molecules signal IL and Toll-like receptors to activate signaling cascades involving Fas ligand, nuclear factor (NF-. kappa.B) and MEKK/JNK pathways. In ICH, anti-inflammatory signaling is activated in a cascade driven by downstream calbindin, cytoskeletal and ribosomal proteins, and c-Myc from annexins a1 and a2, IL-10, and transforming growth factor-beta (TGF- β). ICH down-regulates the parallel neuronal signaling system in the tissue surrounding the hematoma. These down-regulated neuronal genes include molecules that play a role in glutamate signaling, presynaptic structure, postsynaptic structure, and numerous ion channels and calcium signaling proteins. At the cellular level, specific cell types respond to ICH with altered expression of these genes. Astrocytes in the peripheral border of the hematoma express aquaporin 9 and a tissue inhibitor of metalloproteinase-1 (TIMP-1). Annexin a2 is induced in inflammatory cells at bleeding and in neurons located at the margins of the bleeding site. Oligodendrocytes in the damaged white matter express the inflammatory chemokine CCR 1. Inflammatory and endothelial cells immediately surrounding the hematoma and at the hemorrhage site express the IL receptor IL1R 1. At the protein level, the selective member of the molecular cascade induced in the tissue surrounding the hematoma after ICH is localized in these two tissue compartments together with different glial and neuronal cell types.

Thus, ICH induces several biomarkers that can be used to study ICH: (i) inflammatory cytokine signaling centered on tumor necrosis factor (TNF- α), IL-1 β, and IL-6; (ii) glial edema-associated genes including aquaporin 4 and Vascular Endothelial Growth Factor (VEGF); and (iii) the expression of matrix metalloproteinase 9(MMP9) and serum protease (plasminogen) as mediators of secondary extracellular matrix damage.

Tumor necrosis factor (TNF- α, TNF, cachexin, cachectin) is a cytokine involved in systemic inflammation, immune cell regulation, apoptotic death and tumor formation inhibition. Two receptors (TNF-R1 and TNF-R2) bind TNF- α. TNF-R1 is expressed in most tissues, while TNF-R2 is present in cells of the immune system. The binding of TNF- α to the TNF receptor results in a conformational change of the receptor which allows the initiation of several cascades including (i) the activation of NF- κ B (which mediates the transcription of a diverse range of proteins involved in cell survival and proliferation, inflammatory responses and anti-apoptotic factors); (ii) activation of MAPK pathways (including JNK pathways involved in cell differentiation, proliferation and, in general, pro-apoptosis); and (iii) inducing death signaling.

Interleukin (IL-6), a cytokine with pro-and anti-inflammatory properties, is secreted by T cells and macrophages, among other cells, to stimulate an immune response against trauma, and by smooth muscle cells in the media of many blood vessels. IL-6 is an important mediator of the febrile and acute phase reactions. IL-6 signals through the cellular type 1 cytokine receptor (by IL-6Ra (ligand binding chain) and gp130 (signal transduction component). when IL-6 interacts with its receptor, it triggers gp130 and IL-6R proteins to form complexes, thus activating the receptor.

Interleukin 1 β (IL-1 β) is a cytokine produced by activated macrophages as a proprotein that is proteolytically processed into its active form by caspase 1 (CASP/ICE). IL-1 β is an important mediator of the inflammatory response and is involved in several cellular activities, including but not limited to cell proliferation, differentiation and apoptosis.

Aquaporin 4 is an integral membrane protein that directs water across the cell membrane. It is constitutively expressed in the basolateral cell membrane of the major collecting duct cells in the kidney. Aquaporin 4 is also expressed in astrocytes and is upregulated by direct injury to the central nervous system.

Vascular Endothelial Growth Factor (VEGF) stimulates the growth of new blood vessels. All members of the VEGF family stimulate cellular responses by: binds to tyrosine kinase receptors (VEGFR) on the cell surface, causing them to dimerize and become activated (albeit at different sites, times and extents) via transphosphorylation.

Matrix Metalloproteinases (MMPs) are zinc-dependent endopeptidases capable of degrading a variety of extracellular matrix proteins. MMPs share a common domain structure, including a propeptide, a catalytic domain, and a hemagglutinin-like C-terminal domain connected to the catalytic domain by a flexible hinge region. MMPs are known to be involved in the cleavage of cell surface receptors, the release of apoptotic ligands (e.g., FAS ligands), and chemokine/cytokine inactivation/activation. MMPs are generally thought to play a role in cell proliferation, migration (adhesion/dispersion), differentiation, angiogenesis, apoptosis, and host defense.

Requirement of ICH therapy and model System

Spontaneous ICH is twice as common as subarachnoid hemorrhage (SAH) and is more likely to cause death or severe disability than cerebral infarction or SAH. The benefits of surgery or medical treatment of idiopathic ICH patients remain uncertain.

Age-related age and hypertension are significant risk factors for spontaneous ICH. The most important cause of ICH is generally thought to be pathophysiological changes in arterioles and arterioles due to persistent hypertension. In addition, cerebral amyloid angiopathy is increasingly considered to be the cause of cerebral lobe hemorrhage in the elderly. Other causes of ICH include vascular malformations, ruptured aneurysms, blood clotting disorders, use of anticoagulants and thrombolytics, cerebral infarction bleeding, midbrain tumor bleeding, and drug abuse.

Although guidelines for medical treatment and surgical removal of ICH are available, the treatment of ICH by neurologists and neurosurgeons varies enormously around the world. Despite the lack of proven benefits of surgical removal of ICH, it is estimated that over 7,000 such procedures are performed annually in the united states. The major complication is re-bleeding. Postoperative rebleeding occurs in up to 30% of cases and increases morbidity and mortality.

There is an urgent need for new therapeutic agents and new therapeutic methods for preventing bleeding recurrence in ICH. In addition, the lack of an accurate and reproducible model system for ICH has compromised the research and development of potential new therapeutics and new therapeutic approaches to prevent re-bleeding.

4.1. Subdural hematoma (SDH)

Subdural hematoma (SDH) is a form of traumatic brain injury in which blood collects between the dura mater and the arachnoid. The subdural space is a potential cavity formed by the separation of cells in the dural margin cell layer. The outer wall is the dura mater, a layer of poorly vascularized dense fibrous membrane, and the inner wall is the vascularized arachnoid without a capillary bed. The inner layer of the dura mater and the remaining dural margin layer cells have a high reactive potential for cell formation and contain a very fine network of interconnected capillaries. Unlike epidural hematomas, which are typically caused by arterial tears resulting in blood build-up between the dura mater and the skull, subdural bleeding is typically produced by venous tears through the subdural space, causing blood to collect within the inner meningeal layer or the dural margin cell layer of the dura mater. This bleeding often separates the dura from the arachnoid layer.

Subdural hematomas are classified as acute, subacute, and chronic, depending on the rate of their onset.

Acute SDH is usually due to trauma and is the most fatal of all head injuries. They have a high mortality rate if ASDH is not treated promptly by surgical decompression. When an acute hematoma is confined to the subdural space without a arachnoid tear, the hematoma breaks open inside the dural border cell layer.

Subacute SDH is generally described as those hematomas that form within 2 to 6 weeks of head trauma.

Chronic SDH results from bleeding occurring within the "potential" cavity between the dura mater and arachnoid mater and is generally described as requiring about 6 weeks to develop. This "potential" compartment is so described because, under normal circumstances, the brain and its arachnoid and pia mater linings are in direct contact against the dura mater. However, with age, cortical atrophy occurs and a true "subdural space" can develop due to the separation of cells in the dural border cell layer. Subsequently, the small veins that flow out of the cortex may be adhered to the dura mater, and these "bridging" veins traverse the now dilated subdural space. Thus, these "bridging" veins are prone to tearing and bleeding due to any trauma that will exert inertial forces on the brain. This may occur after a relatively minor accident, such as the elderly slipping or falling, a head bump into the doorway, or some other relatively minor accident. Acute bleeding is usually stopped when a torn vein coagulates or when the pressure created by the enlarged blood clot exceeds the pressure of the bleeding vein. Bleeding can recur with other trauma or for reasons not clearly understood, and the clot itself can expand in size within the following weeks following the initial accident.

4.2. Pathogenesis of chronic SDH

The pathogenesis of chronic SDH has been controversial for over a century and is still under speculation. The most prevailing theory involves recurrent bleeding from the haematoma sac and associated excessive fibrinolysis. It is generally believed that chronic SDH is formed in the dural border cell layer of the haematoma lumen which then continues to form the characteristic outer and inner membranes. Although there are a few blood vessels in the intima, which originates primarily from the arachnoid, the adventitia contains many fragile large capillary vessels (also known as "sinus-like vessels") that are often the origin of repeated multifocal hemorrhages. It is considered that such repeated bleeding from the adventitia is a causative factor of progressive expansion of hematoma.

Chronic SDH is often accompanied by increased fibrinolytic activity that destabilizes hemostatic clots, leading to recurrent bleeding and hematoma cysts.

Generally, the surface of the serous cavity within the body absorbs any foreign material when contacted. Although the subdural space is not completely analogous to the serous space elsewhere in the body, it is generally thought to behave similarly, and thus, the accumulation of blood, fibrin and Fibrin Degradation Products (FDPs) within the subdural space can lead to cell formation with resorption of subdural aggregates or to the formation of progressively expanding SDH. Low brain back pressure, excessive subdural aggregates or physiological brain atrophy may be a slow, progressive expansion of chronic SDH's. Dural border cells typically form hematomas, proliferate and produce pseudomembranes (adventitia and intima) during or later in the second cycle, and eventually the hematomas are intersected by collagen and elastic fibers and sprouting capillaries (sinus-like vessels). These vessels are fragile and are known to bleed easily. The inner surface of the hematoma forms its own pseudomembrane, separating the blood clot from the arachnoid. Thus, the pseudomembrane remains exceptionally vulnerable to traction as long as the proliferative changes continue. Although the increase in collagen will strengthen the pseudomembrane and culminate in fibrotic healing of the lesion, a vicious circle can develop in which the microscopic trauma and fibrinolytic activity in the hematoma fluid trigger further proliferation of dural border layer cells and bleeding from the delicate sinus-like neovessels in the outer membrane, resulting in more pseudomembrane formation, SDH bleeding and expansion of SDH.

Histological studies of the outer membrane of hematomas have demonstrated the enormous proliferative potential and fragility of numerous large capillaries. The most characteristic clinical pathology aspect of the adventitia of chronic SDH seems to be at best its predisposition to recurrent, multifocal hemorrhage of large capillary angiogenesis. The general features of endothelial cells of large capillaries are large lumens, attenuated or flattened endothelial cells, lack of cytoplasmic intercalation, less tight cell junctions, gap junctions and a smaller or absent basement membrane thickness. These traits indicate that large capillaries are quite fragile, prone to bleeding and result in abnormally high vascular permeability. The number and extent of gaps between endothelial cells ranging in size from 0.6 μm to 8 μm suggests that they may be responsible for most of the leakage into the adventitial tissue and the hematoma lumen. Several studies have shown that adjacent endothelial cells can temporarily separate, allowing red blood cells as well as plasma to escape into the lumen of large capillaries. The mechanisms by which endothelial spaces form are not completely understood; increased intraluminal hydrostatic pressure or endothelial contraction may induce adjacent endothelial cells to separate, and/or perivascular leakage of blood material from large capillary vessels with such endothelial spaces and incomplete basement membranes may contribute to the expansion of chronic SDH.

In addition, several studies have shown that the experimentally induced growth content of chronic SDH is directly proportional to the thickness of the large capillary layer and the extent of leakage. Several studies have shown that the pathogenesis of chronic SDH expansion is caused by direct bleeding of large capillary vessels, infiltration of perisinus edema fluid into the hematoma cavity, and/or disruption of the hemorrhagic cavity formed in the adventitia.

4.3. Classification of Chronic SDH

Several classification schemes for chronic SDH have been proposed. Nomura (Nomura, S. et al, J. Neurosurg.81: 910-. Nakaguchi (Nakaguchi, H. et al, J.Neurosurg.95: 256-charge 62,2001) defines four neuroimaging classes of hematomas based on CT scan results; 1) a uniform density pattern; 2) stratified, defined as a uniform density subtype, accompanied by a high density layer along the intima; 3) a layered or separated type, comprising two portions with different densities, between which a boundary exists; and 4) trabecular density type, in which a high density separator between an inner membrane and an outer membrane is lined with a low to iso-density background. Frati (j. neurosurg.100:24-32,2004) proposed a combination of the Nomura classification scheme and the Nakaguchi classification scheme that divided hematomas into 4 different classes: class 1, isolated or stratified; category 2, hierarchical or hybrid; class 3, trabecular (classified by Nomura into mixed density hematoma class); and class 4, high, low, or iso-density type described by Nomura, which is also defined by Nakaguchi as a uniform density type.

4.4. Biomarkers for chronic SDH

Several studies have suggested that the pathophysiology of chronic SDH is similar to that of inflammatory responses. According to one theory, after trauma and once the subdural space has been created, CSF or blood collects inside the dural limbal cell layer. Blood aggregates may be caused by tears in the bridging vein after trauma. In elderly patients, as a result of brain atrophy, the dural border cell layer and bridging veins stretch and can be easily damaged by traumatic events. Once this epidural space has been created, the cells in the dural border layer begin to proliferate, which represents the first step in the pathogenesis of chronic SDH. Mesenchymal cells proliferate, differentiate and form an outer or adventitial membrane (an inflammatory sac or membrane) around a blood clot or CSF. The outer membrane of chronic SDH consists of granulation tissue in which several types of inflammatory cells (mast cells, eosinophils, neutrophils, monocytes, macrophages, endothelial cells and fibroblasts) are activated and recruited continuously. This membrane also contains immature blood vessels and connective tissue fibers and generally constitutes a source of inflammatory, angiogenic, fibrinolytic and clotting factors.

Immunohistochemical analysis has shown the expression of the cytokine VEGF in inflammatory cells infiltrating the pseudomembrane of chronic SDH, mainly in plasma cells and tissue macrophages. Some studies that have examined the role of VEGF in angiogenesis and vascular hyperpermeability suggest that inflammation is responsible for adventitial angiogenesis (meaning the physiological process involved in the growth of new blood vessels from pre-existing blood vessels). Thus, after trauma, a series of events in the natural process of chronic SDH include local inflammation, angiogenesis, vascular leakage or permeability (due to immature new blood vessels), bleeding, hypercoagulative activity, hyper fibrinolytic activity and sustained vascular permeability (due to bradykinin, which is activated by plasmin from high molecular weight kininogens). This leads to further inflammation caused by proinflammatory factors such as cytokines and bradykinin release, producing a self-enhancing vicious cycle that causes frequent rebleeding and expansion of chronic SDH. Thus, some have suggested that the biomarkers IL-6 and IL-8 may be suitable for the study of chronic SDH.

In general, IL-6 and IL-8 are markers of inflammatory processes. IL-6 and IL-8 are produced by many different cell types, including stimulated monocytes, macrophages, fibroblasts, endothelial cells, T cells, B lymphocytes, granulocytes, smooth muscle cells, eosinophils, chondrocytes, osteoblasts, mast cells, and glial cells.

Briefly, IL-6 affects one of the major physiological mediators of immune and inflammatory responses and acute phase responses. Inflammation within the nervous system, very high levels of IL-6 can often be observed in the CSF of patients with bacterial or viral meningitis, as well as in patients with gliomas. The role of IL-8 in inflammatory processes is well established. IL-8 differs from all other cytokines in its specific ability to enhance the affinity of adhesion molecules for neutrophils, activate neutrophils and mediate neutrophil chemotaxis. In addition, IL-8 supports angiogenesis and can play a role in angiogenic processes such as granulation tissue, wound healing and tumor growth.

Chronic SDH has in general some characteristics of a chronic inflammatory process. In hematomas, IL-6 and IL-8 are secreted by fibroblasts and by endothelial cells and inflammatory cells that infiltrate the outer membrane. IL-6 and IL-8 production is increased by proinflammatory factors such as platelet activating factor, bradykinin and thrombin released after bleeding.

4.5. Chronic SDH treatment and model System requirements

Putative risk factors for chronic SDH include age; alcoholism; medical disorders such as liver dysfunction, kidney disease, diabetes, dementia or coagulopathy; hemodialysis; use of anticoagulants, antiplatelet agents or chemotherapeutic agents; presence of cerebrospinal fluid shunt or treated hydrocephalus; draining cerebrospinal fluid after surgery; and/or other causes of a decrease in brain size relative to the fixed size of the skull. The appearance of chronic SDH on CT or Magnetic Resonance Images (MRI) (stratified hematomas, among other types) and the methods of treating chronic SDH (drilling, craniotomy, and trephine with or without irrigation) can impact the development of chronic SDH. Chronic SDH is a common disease in the elderly and its incidence is highest in people over 65 years of age (58 cases per 100,000 individuals). In about 60% to 80% of cases, mildly traumatic events are reported with bleeding first; however, sometimes a slightly traumatic attack may not be detectable. These putative risk factors of recurrence have been discussed by way of illustration in several reports where the results of controversial studies are not uncommon.

Surgery is a treatment option for chronic SDH. Several different forms of surgery have been used: craniotomies, drilling with or without irrigation and/or a closed drainage system, and trephine trephines that directly access the hematoma at the site of greatest thickness of the hematoma. However, the symbiotic disease associated with chronic SDH can impair its prognosis and the outcome of surgical medication. In addition, the recurrence rate of chronic SDH after surgery is between 3.7% and 30%. Recurrence increases the probability of mortality and morbidity due to chronic SDH.

The lack of an accurate and reproducible model system for chronic SDH has compromised the work of studying and developing new therapeutics and new therapeutic approaches to prevent the recurrence of chronic SDH. Existing attempts to develop animal models of chronic SDH have exhibited significant limitations including, but not limited to, lack of reproducibility.

5. Blood coagulation

Hemostasis is the cessation of blood loss from a damaged blood vessel. Platelets first adhere to macromolecules in the subendothelial region of the damaged vessel; they subsequently aggregate to form primary hemostatic plugs. Platelets stimulate local activation of plasma clotting factors, resulting in the production of fibrin clots that strengthen the platelet aggregates. Later, as wound healing occurs, platelet aggregates and fibrin clots degrade (Goodman & Gilman's The pharmaceutical Basis of Therapeutics, joelg.hardman and Lee e.limbed (eds.), McGraw-Hill,2001, pages 1519-20).

Coagulation involves a series of zymogen activation reactions. At each stage, the precursor protein or zymogen is converted to an active protease by cleavage of one or more peptide bonds in the precursor molecule. The components that may be involved in each stage include proteases from the previous stage, proenzymes, non-enzymatic protein cofactors, calcium ions and the forming surface provided by damaged blood vessels and platelets in vivo. The final protease to be produced is thrombin (factor IIa).

Fibrinogen is a 330,000 dalton protein consisting of three pairs of polypeptide chains (designated α, β, and γ) covalently linked by disulfide bonds. Thrombin converts fibrinogen to fibrin monomer (factor IA) by cleaving the fibrinopeptides a (16 amino acid residues) and B (14 amino acid residues) from the amino termini of the alpha and beta chains, respectively. Removal of the fibrinopeptides allows the fibrin monomers to form a gel. Initially, fibrin monomers are non-covalently bound to each other. Subsequently, factor XIIIa catalyzes an interchain transglutaminase reaction that crosslinks adjacent fibrin monomers to enhance clot strength.

Fibrin is involved in the activation of thrombin-activated factor XIII and plasminogen activator (t-PA). Fibrin specifically binds activated coagulation factors, factor Xa and thrombin and embeds them within the fibrous network, thus acting as temporary inhibitors of these enzymes, which are still active and can be released during fibrinolysis. Recent studies have shown that fibrin plays an important role in the inflammatory response.

Protease zymogens involved in blood coagulation include factor II (prothrombin), VII, IX, X, XI, XII and prekallikrein. Factor V and factor VIII are homologous proteins of 350,000 daltons. Factor VIII circulates in plasma, binding to von willebrand factor, while factor V is present both free in plasma and as a component of platelets. Thrombin cleaves V and VIII to produce activated factors (Va and VIIIa) which have at least 50 times the coagulant activity of the precursor form. Factors Va and VIIIa themselves have no enzymatic activity, but act as cofactors for the proteolytic efficiency of Xa and IXa, respectively. Tissue Factor (TF) is a non-enzymatic lipoprotein cofactor that greatly increases the proteolytic efficiency of VIIa. It is present on surfaces that are not normally in contact with plasma cells (e.g., fibroblasts and smooth muscle cells) and initiates coagulation outside of damaged blood vessels.

Two coagulation pathways were identified: the intrinsic coagulation pathway (so called because all components are inherent to the plasma) and the extrinsic coagulation pathway. Exogenous and endogenous systems converge to activate the final common pathway responsible for fibrin formation. Figure 6 shows a schematic of a classical coagulation cascade. These systems are generally thought to function together and interact in vivo.

The exogenous system (tissue factor pathway) produces a thrombin surge and is initiated when tissue thromboplastin activates factor VII. Once the blood vessel is damaged, TF is exposed to blood and the circulating enzyme factor VII (pretransformin) in the blood. Once bound to TF, FVII is activated to FVIIa by different proteases including thrombin (factor IIa), factor Xa, IXa, Xlla and the FVIIa-TF complex itself. The FVIIa-TF complex activates factor IX and factor X. FXa activation by FVIIa-TF is almost immediately inhibited by Tissue Factor Pathway Inhibitor (TFPI). Factor Xa and its cofactor Va form a prothrombin complex which activates the conversion of prothrombin to thrombin. Thrombin then activates other components of this coagulation cascade, including FV and FVIII (which activates FXI, which in turn activates FIX), and FVIII is activated and released from binding to vWF (von Willebrand factor). FVIIa and FIXa together form a "tenase" complex which activates FX and the cycle continues as such.

The endogenous system (contact activation pathway) is initiated when blood contacts any surface except normal endothelial cells and blood cells. The endogenous system begins with the formation of a primary complex on collagen by High Molecular Weight Kininogen (HMWK), prekallikrein and FXII (Hageman factor). Prekallikrein is converted to kallikrein and FXII becomes FXIIa. FXIIa converts FXI to FXIa. Factor XIa activates FIX, which together with its cofactor FVIIIa forms the tenase complex, which activates FX to FXa.

As is currently understood, in vivo coagulation is a 3-step process centered on the cell surface. FIG. 7 shows a schematic representation of a cell surface based in vivo coagulation model (Monoe Arterioscler Thromb Vase biol. 2002; 22: 1381-. In the first step, coagulation is initiated primarily by the use of tissue factors that, when activated by inflammation, are present on the subendothelial membrane (tissue that is not normally exposed to blood, activated monocytes, and endothelium). Factor VII and factor VIIa bind to tissue factor and adjacent collagen. The factor VIIa-tissue factor complex activates factor X and factor IX. Factor Xa activates factor V, forming a prothrombin complex (factor Xa, factor Va and calcium) on cells expressing tissue factor. In the second step, the coagulation effect is amplified when platelets adhere to the site of injury in the blood vessel. Thrombin is activated by platelet adhesion and then acts to sufficiently activate platelets, enhance their adhesion, and release factor V from platelet alpha particles. Thrombin on the surface of activated platelets activates factors V, VIII and XI, followed by factor IX. the tenase complex (factors IXa, VIIIa and calcium) is now present on platelets where factor Xa can be produced, and another prothrombinase complex can be produced on platelets, allowing large-scale production of thrombin. Proliferation is the third step and is a combination of prothrombinase complex activation that allows for the production of large amounts of thrombin from prothrombin. More platelets can be recruited, as well as fibrin polymer and factor XIII activation.

Natural anticoagulant mechanism

Platelet activation and coagulation do not normally occur intravascularly. Thrombosis (meaning the pathological process in which platelets aggregate and/or fibrin clots occlude blood vessels) is prevented by several regulatory mechanisms that require normal vascular endothelium. Prostacyclin (PGI2), an arachidonic acid metabolite synthesized by endothelial cells, inhibits platelet aggregation and secretion. Antithrombin is a plasma protein that inhibits coagulation factors of both the intrinsic and common pathways. Heparan sulfate proteoglycans synthesized by endothelial cells stimulate antithrombin activity. Protein C is a plasminogen that is homologous to factors II, VII, IX and X. Activated protein C in combination with its non-enzymatic cofactor (protein S) degrades cofactors Va and VIIIa and thereby greatly reduces the rate of activation of prothrombin and factor X. Protein C is activated by thrombin only in the presence of thrombomodulin, a membrane-integrated membrane protein of endothelial cells. Like antithrombin, protein C appears to produce an anticoagulant effect in the vicinity of intact endothelial cells. Tissue Factor Pathway Inhibitor (TFPI), which is present in the lipoprotein fraction of plasma, inhibits the factor Xa and factor VIIa-tissue factor complex when bound to factor Xa.

Fibrinolysis

The degradation of fibrin is called "fibrinolysis". The fibrinolytic system dissolves intravascular clots by the action of plasmin, an enzyme that digests fibrin. Plasminogen, an inactive precursor, is converted to plasmin by cleavage of a single peptide bond. Plasminogen (EC 3.4.21.7; PLG) degrades many blood plasma proteins, including fibrin clots.

The fibrinolytic system is regulated to remove unwanted fibrin thrombi, while fibrin in the wound remains to maintain hemostasis. FIG. 8 is a schematic representation of the fibrinolytic pathway (Meltzer et al, SeminsThrombosis Hemostasis 2009,35: 469-77). In response to a variety of signals, including stasis due to vascular occlusion, the serine protease tissue plasminogen activator (t-PA) is released from endothelial cells. t-PA is rapidly cleared from the blood or inhibited by circulating inhibitors (plasminogen activator inhibitor-1 and plasminogen activator inhibitor-2) and thus has a minor effect on circulating plasminogen. tPA binds to fibrin and converts plasminogen, which also binds to fibrin, to plasmin. Plasminogen and plasmin bind to fibrin at a binding site located near the N-terminus of its lysine-rich (Lys, K) residues. These sites are also needed for plasmin binding to the inhibitor α 2-antiplasmin. Alpha 2-antiplasmin forms a stable complex with plasmin, thereby inactivating plasmin. Thus, plasmin binding fibrin is protected from inhibition (Goodman & Gilman's the pharmacological Basis of Therapeutics, Joel G.Hardman and Lee E.Limbird (eds.), McGraw-Hill,2001, pages 1531-32).

Plasminogen contains a high affinity, amino-terminal lysine-containing binding site that mediates binding of plasminogen (or plasmin) to fibrin; this enhances fibrinolysis. These sites are located in the amino-terminal secondary structural motif (called "kringles") that specifically binds to lysine and arginine residues of fibrin (ogen); when converting from plasminogen to plasmin, plasmin acts as a serine protease by cutting the chain at the C-terminus of amino acids to these lysine and arginine residues. Thus, plasmin action on the clot initially produces a cut in the fibrin and further digestion leads to solubilization. These sites also promote plasmin complex formation with α 2-antiplasmin (the major physiological plasmin inhibitor).

6. Anti-fibrinolytic agents

"anti-fibrinolytic agents" (meaning drugs used to prevent the dissolution of fibrin clots) have been used to treat some hemorrhagic conditions. These drugs (typically lysine analogs) are potent inhibitors of enzymes involved in the fibrinolytic pathway. Aprotinin (bovine trypsin inhibitor, or BPTI, commercially available as Trasylol) that has been used to reduce bleeding during surgery when administered to humans^TMSold) (a bovine protein that inhibits plasmin) is associated with a fatal allergic response.

6.1. Amino caproic acid

Amistar binds reversibly to the tricyclic domain of plasminogen (the autoproteins domain folded into a large loop stabilized by 3 disulfide bonds). By this binding, it prevents activation of plasminogen by its activator to plasmin and prevents plasmin from acting on fibrin.

Amistar is used to enhance hemostasis when fibrinolysis contributes to bleeding. Fibrinolytic bleeding can often be accompanied by surgical complications following cardiac and other types of surgery (with or without cardiac bypass) and portal bypass; hematological disorders such as megakaryocytic thrombocytopenia (with aplastic anemia); acute and life threatening pre-placentas; cirrhosis of the liver; rupture of intracranial aneurysm; intracerebral and SDH; and neoplastic diseases such as cancer of the prostate, lung, stomach and cervix. Urofibrinolysis (generally a normal physiological phenomenon) can contribute to excessive fibrinolytic bleeding of the urinary tract with surgical hematuria (after prostatectomy and nephrectomy) or non-surgical hematuria (with polycystic or neoplastic disease of the genitourinary system).

Amica inhibits the action of plasminogen activator and to a lesser extent plasmin activity. Amistar inhibits fibrinolysis which could theoretically lead to coagulation or thrombosis. Several studies have shown an increased incidence of certain neurological deficits (e.g., hydrocephalus, cerebral ischemia, or cerebral vasospasm) associated with the use of antifibrinolytic agents in the treatment of subarachnoid hemorrhage (SAH). The drug association is still unclear, but one theory suggests that these complications are due to promotion of thrombosis due to these drugs. Therefore, it is suggested that amica should not be administered with factor IX complex concentrates or with anti-inhibitor coagulation concentrates, as the risk of thrombosis may increase.

6.2. Tranexamic acid

Tranexamic acid(a synthetic derivative of lysine) is a competitive inhibitor of plasminogen activation and, at much higher concentrations, is a noncompetitive inhibitor of plasmin.

It is about 10 times stronger in vitro than amica. Tranexamic acid binds more strongly to the strong and weak receptor sites of the plasminogen molecule than amicor at a ratio corresponding to the difference in potency between these compounds.

Tranexamic acid is used as a first-line non-hormonal therapy for dysfunctional uterine bleeding, severe bleeding associated with uterine fibroids and as a second-line therapy for factor VIII in hemophilia.

6.3. Factor VII

Recombinantly activated factor VIIa is approved for hemophilia (along with inhibitors) in north america and europe. Factor VII acts locally at the site of tissue damage and vessel wall destruction by binding to tissue factor, thus producing a small amount of thrombin sufficient to activate platelets. At pharmacological doses, factor VII directly activates factor X on the surface of activated platelets, leading to an acceleration of thrombin burst and coagulation (Mayer, S. et al, NEJM,2008,358: 2127-.

6.4. Aprotinin

Aprotinin, a bovine protein consisting of 58 amino acid residues with a molecular weight of 6,512 daltons, was isolated in the 30 s of the twentieth century because of kallikrein and trypsin inhibition and was used clinically since 1959. By inhibiting fibrinolysis and preserving platelet function, aprotinin has been shown to reduce blood loss and transfusion requirements in cardiac surgery, lung transplantation, and liver transplantation and hip replacement surgery. Other disease-related indications are excessive fibrinolytic hemostatic disorders and complications of thrombolytic therapy. The most common commercial preparation is Trasylol^TM(Bayer AG, Leverkusen, Germany) and Antagosan^TM(Aventis Pharma, Frarikfurt/M, Germany) (Beierlein, W. et al, Ann. Thorac. Surg.,2005,79: 741-748).

Due to its anti-fibrinolytic effect, aprotinin is added to the fibrin sealant in order to achieve hemostasis, even when the fibrinolytic activity is increased. Ready-to-use fibrin sealant kits have been marketed in europe since 1974 and in the us since 1998. The most common kit is Beriplast^TM(Centeon, horse)Erberg, Germany), Tissucol/Tisseel^TM(Baxter Hyland Immuno Division, Vienna, Austria) and Hemaseel^TM(Hemacure, Montreal, Canada). Tachocomb^TM(Nycomed, Roskilde, Denmark), a hemostatic solid equine collagen fleece, also contains small amounts of aprotinin (Beierlein, W. et al, Ann. Thorac. Surg.,2005,79: 741-748).

7. Drug delivery from bioresorbable polymers

The combination of a biodegradable polymer with a drug or pharmaceutically active compound may allow for a formulation capable of sustained release of the drug when injected or inserted into the body.

Site-specific activity is generally effective if the site into which the in vivo formulation is deposited is a fluid-filled cavity or some type of cavity, e.g., the subarachnoid space, the subdural space of chronic SDH, or the cavity left after surgical removal of hematomas, tumors, or vascular malformations in the brain. This provides a high concentration of drug at the site where activity is required and a lower concentration in the rest of the body, thus reducing the risk of unwanted systemic side effects.

Site-specific delivery systems include, for example, the use of microparticles (about 1 μm to about 100 μm in diameter), thermoreversible gels (e.g., PGA/PEG), and biodegradable polymers (e.g., PLA, PLGA) that can be in the form of thin films.

The delivery characteristics of the drug and the in vivo degradation of the polymer may also be modified. For example, polymer conjugation can be used to alter circulation of a drug within the body and to achieve tissue targeting, reduce irritation, and improve drug stability.

Despite all these possibilities, no one has applied such polymers to deliver therapeutic agents locally within the human brain to treat hemorrhagic brain disease.

The invention provides therapeutic compositions, methods for identifying therapeutic agents to treat a hemorrhagic condition of the brain, methods for treating a hemorrhagic condition of the brain, and accurate and repeatable model systems for chronic SDH and ICH.

Brief description of the invention

According to one aspect, the invention provides a method for treating hematoma expansion or recurrent rebleeding resulting from a hemorrhagic condition in the brain, the method comprising: (a) providing a pharmaceutical composition comprising (i) a therapeutically effective amount of an anti-fibrinolytic agent and (ii) a pharmaceutically acceptable carrier; (b) administering such a pharmaceutical composition into or at a distance proximal to an intracerebral hematoma; and (c) improving patient outcome. According to one embodiment, the hemorrhagic condition results from Traumatic Brain Injury (TBI). According to another embodiment, the hemorrhagic condition is post-surgical removal of a hematoma followed by hemorrhage. According to another embodiment, the hemorrhagic condition is a chronic subdural hematoma. According to another embodiment, the hemorrhagic condition is an intracerebral hematoma. According to another embodiment, the intracerebral hematoma is a spontaneous intracerebral hematoma. According to another embodiment, the intracerebral hematoma is a traumatic intracerebral hematoma. According to another embodiment, the hemorrhagic condition is rebleeding after craniotomy. According to another embodiment, craniotomy procedures are performed to treat brain cancer. According to another embodiment, a craniotomy procedure is performed to treat an intracerebral vascular abnormality. According to another embodiment, a craniotomy procedure is performed to treat a cerebral aneurysm. According to another embodiment, the administration is implantation. According to another embodiment, the anti-fibrinolytic agent is-aminocaproic acid (amistar). According to another embodiment, the anti-fibrinolytic agent is factor VII. According to another embodiment, the factor VII is a recombinant factor VII. According to another embodiment, the anti-fibrinolytic agent is tranexamic acid. According to another embodiment, the anti-fibrinolytic agent is aprotinin. According to another embodiment, the pharmaceutically acceptable carrier is a controlled release carrier. According to another embodiment, the pharmaceutically acceptable carrier is a sustained release carrier. According to another embodiment, the anti-fibrinolytic agent is embedded in the sustained-release carrier. According to another embodiment, the anti-fibrinolytic agent is coated on the sustained-release carrier. According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration. According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration. According to another embodiment the sustained release carrier is a microparticle. According to another embodiment, the sustained release carrier is a nanoparticle. According to another embodiment, the sustained release carrier comprises a biodegradable polymer. According to another embodiment, the biodegradable polymer is a synthetic polymer. According to another embodiment, the biodegradable polymer is a naturally occurring polymer. According to another embodiment, the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof. According to another embodiment, the synthetic polymer is polyglycolic acid (PGA). According to another embodiment, the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA) or polycaprolactone. According to another embodiment, the sustained release carrier is a hydrogel. According to another embodiment, the naturally occurring biopolymer is a protein polymer. According to another embodiment, the naturally occurring polymer comprises hyaluronic acid. According to another embodiment, the naturally occurring polymer comprises less than 2.3% hyaluronic acid. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 10 mm. According to another embodiment, the pharmaceutical composition exhibits a localized pharmacological effect. According to another embodiment, the pharmaceutical composition exhibits a pharmacological effect through the brain.

According to another aspect, the invention provides a site-specific, sustained-release pharmaceutical composition for treating hematoma expansion or recurrent rebleeding resulting from a hemorrhagic condition in the brain, the pharmaceutical composition comprising: (a) a therapeutically effective amount of an anti-fibrinolytic agent and (b) a pharmaceutically acceptable carrier, wherein the pharmaceutically acceptable carrier is a sustained release carrier. According to another embodiment, the hemorrhagic condition is rebleeding following traumatic brain injury. According to another embodiment, the hemorrhagic condition is a chronic subdural hematoma. According to another embodiment, the hemorrhagic condition is an intracerebral hematoma. According to another embodiment, the intracerebral hematoma is a spontaneous intracerebral hematoma. According to another embodiment, the intracerebral hematoma is a traumatic intracerebral hematoma. According to another embodiment, the hemorrhagic condition is rebleeding after craniotomy. According to another embodiment, craniotomy procedures are performed to treat brain cancer. According to another embodiment, a craniotomy procedure is performed to treat an intracerebral vascular abnormality. According to another embodiment, a craniotomy procedure is performed to treat a cerebral aneurysm. According to another embodiment, the anti-fibrinolytic agent is-aminocaproic acid (amistar). According to another embodiment, the anti-fibrinolytic agent is factor VII. According to another embodiment, the factor VII is a recombinant factor VII. According to another embodiment, the anti-fibrinolytic agent is tranexamic acid. According to another embodiment, the anti-fibrinolytic agent is aprotinin. According to another embodiment, the pharmaceutically acceptable carrier is a controlled release carrier. According to another embodiment, the pharmaceutically acceptable carrier is a sustained release carrier. According to another embodiment, the anti-fibrinolytic agent is embedded in the sustained-release carrier. According to another embodiment, the anti-fibrinolytic agent is coated on the sustained-release carrier. According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration. According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration. According to another embodiment, the sustained release carrier comprises microparticles. According to another embodiment, the sustained release carrier comprises nanoparticles. According to another embodiment, the sustained release carrier comprises a biodegradable polymer. According to another embodiment, the biodegradable polymer is a synthetic polymer. According to another embodiment, the biodegradable polymer is a naturally occurring polymer. According to another embodiment, the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof. According to another embodiment, the synthetic polymer is polyglycolic acid (PGA). According to another embodiment, the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA) or polycaprolactone. According to another embodiment, the sustained release carrier is a hydrogel. According to another embodiment, the naturally occurring polymer is a protein polymer. According to another embodiment, the protein polymer is synthesized from a self-assembled protein polymer. According to another embodiment, the naturally occurring polymer is a naturally occurring polysaccharide. According to another embodiment, the naturally occurring polymer comprises hyaluronic acid. According to another embodiment, the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

Drawings

FIG. 1 shows a diagrammatic side view of the human brain (Stedman's Medical Dictionary, 27 th edition, plate 7at A7 (2000).

FIG. 2 shows a diagrammatic sagittal view of the human brain (relative Neuroanatomy & functional neurology, 18 th edition, page 46 (1982)).

Fig. 3 shows a representation of a section of intact meninges from the inner surface of the skull (upper) to the outer surface of the brain (lower). Collagen is present in the periosteal and meningeal layers (large dots, not shown in the direction of the fibrils) and in the subarachnoid space (SAS), usually in the folds of trabecular cells. The dural border cell layer is devoid of extracellular collagen, with few cell junctions, an expanded extracellular space (but without basement membrane) and fibroblasts different from those of the epidural portion. The arachnoid barrier cell layer is essentially free of extracellular space, has numerous cell junctions, is more voluminous in appearance and has a more continuous basement membrane on the surface facing the SAS. Note the continuity of the cell layer from the arachnoid to the dura mater (no intervening space), the characteristic appearance of the arachnoid trabeculae and the relationship of the pia mater (from Haines DE: On the diagnostic of subdural space. Ant Rec 230:3-21,1991).

FIG. 4 shows a diagrammatic view of the Willis loop and major arteries of the brain (relative Neuroanatomy & Functional Neurology, 18 th edition, page 48 (1982)).

Fig. 5 shows an illustrative view of the arterial supply to the cerebral cortex. 1: the orbito-frontal artery; 2: the anterior Rolando artery; 3: the Rolando artery; 4: the parietal anterior artery; 5: parietal posterior artery; 6: the inner canthus artery; 7: the posterior temporal artery; 8: the anterior temporal artery; 9: the orbital artery; 10: the frontal polar artery; 11: arteries at the corpus callosum border; 12: the posterior internal artery; 13: peripheral arteries (coronary and Functional Neurology, 18 th edition, page 50 (1982)).

Fig. 6 is an illustrative flow of the coagulation cascade.

FIG. 7 is a schematic representation of a cell surface based in vivo coagulation model (Monoe Arterioscler Thromb VaseBiol.2002; 22: 1381-1389).

FIG. 8 is a schematic scheme of the fibrinolytic pathway (Meltzer, S.Thrombosiss Hemostasis 2009,35: 469-77).

FIG. 9 shows the histology of subdural hematomas in a mouse model of chronic subdural hematomas formed by a single injection of 6-aminonicotinamide (25mg/kg body weight).

Detailed Description

Vocabulary and phrases

Anatomical terms:

when referring to an animal (which typically has one end with a head and mouth and an opposite end often with an anus and tail), the head end is referred to as the cranial end and the tail end is referred to as the caudal end. Inside the head itself, the beak refers to the direction towards the tip of the nose and the tail to the tail direction. The normally upwardly oriented animal body surface or side drawn away from gravity is the dorsal side; the contralateral side, the side that is usually closest to the ground when walking, swimming or flying, depending on all legs, is the ventral side. On a limb or other appendage, the point closer to the main body is "proximal"; points that are further away are "distal". Three basic reference surfaces were used in the animal anatomy. The "sagittal" plane divides the body into left and right parts. The "midsagittal" plane is midline, i.e., it passes through a midline structure such as the spine, and all other sagittal planes are parallel thereto. The "coronal" plane divides the body into a dorsal part and an abdominal part. The "transverse" plane divides the body into a cranial portion and a caudal portion.

In referring to humans, the body and parts thereof are described always using the assumption that the body is upright. The body parts closer to the head end are "upper" (corresponding to the cranium in animals), while those further away are "lower" (corresponding to the tail in animals). An object near the front of the body is called "anterior" (in animals corresponding to the abdomen); while those near the back of the body are called "posterior" (corresponding to the back in animals). The transverse, axial or horizontal plane is the X-Y plane, parallel to the ground, which separates the upper and lower/foot portions. The coronal or frontal plane is the Y-Z plane, perpendicular to the ground, which separates the anterior and posterior portions. The sagittal plane is the X-Z plane, perpendicular to the ground and coronal planes, which separates the left and right. The median sagittal plane is the particular sagittal plane located just in the center of the body.

Structures near the midline are referred to as medial, and those near the sides of the animal are referred to as lateral. Thus, the medial structures are closer to the median sagittal plane, and the lateral structures are further from the median sagittal plane. Structures within the midline of the body are neutral. For example, the tip of the nose of a human subject is within the midline.

Ipsilateral means on the same side, contralateral means on the other side and bilateral means on both sides. Structures closer to the center of the body are proximal or central, while structures further away are distal or peripheral. For example, the palm is at the distal end of the arm and the shoulder is at the proximal end.

The term "active" as used herein refers to having a pharmacological or biological activity or effect. The term "active ingredient" ("AI", "active pharmaceutical ingredient", "API" or "bulk active") is a substance in a medicament that is pharmaceutically active. As used herein, the phrase "additional active ingredient" refers to a substance that produces a pharmacological activity or any other beneficial activity in addition to the compounds of the composition.

The term "administering" as used herein means administering or applying. The term "administering" as used herein includes in vivo administration as well as direct administration to ex vivo (ex vivo) tissue. Generally, the compositions can be administered orally, buccally, parenterally, topically, by inhalation or insufflation (i.e., oral or nasal inhalation or insufflation), or rectally, systemically, in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants, and vehicles (vehicles), as desired, or can be administered topically or parenterally, such as, but not limited to, injection, implantation, transplantation, topical application.

The term "agonist" as used herein refers to a chemical substance capable of activating a receptor to induce a full or partial pharmacological response. Receptors can be activated or inactivated by endogenous or exogenous agonists and antagonists, resulting in stimulation or inhibition of biological responses. Physiological agonists are substances that produce the same physical response, but do not bind to the same receptor. Endogenous agonists of a particular receptor are compounds naturally produced by the body that bind to and activate the receptor. A superagonist is a compound that is capable of producing a greater maximal response than an endogenous agonist of the target receptor and therefore produces an efficiency of greater than 100%. This does not actually mean that the compound is more potent than the endogenous agonist, but rather a comparison of the maximum possible response that can be produced intracellularly upon receptor binding. Full agonists bind to and activate the receptor and exhibit full efficacy at this receptor. Partial agonists also bind to and activate a given receptor, but have only partial efficacy at that receptor relative to full agonists. Inverse agonists are substances that bind to the same receptor binding site as agonists of the receptor and reverse the constitutive activity of the receptor. Inverse agonists produce the opposite pharmacological effect of the receptor agonist. An irreversible agonist is an agonist that binds permanently to a receptor in such a way that the receptor is permanently activated. It differs from the common agonists in that the binding of an agonist to a receptor is reversible, whereas the binding of an irreversible agonist to a receptor is considered irreversible. This allows a short burst of agonist activity of the compound followed by receptor desensitization and internalization, which in the case of long-term treatment produces a more antagonist-like effect. Selective agonists are specific for a particular type of receptor.

The term "allogeneic" as used herein means genetically different, although belonging to or obtained from the same species.

As used herein, the term "analog" refers to a compound that has a structure similar to another compound, but is different from the compound, e.g., has one or more atoms, functional groups, or substructures.

The term "antagonist" as used herein refers to a substance that counteracts the effect of another substance.

As used herein, the term "antibody" includes, for example, naturally occurring and non-naturally occurring antibodies. In particular, the term "antibody" includes polyclonal and monoclonal antibodies, and fragments thereof. In addition, the term "antibody" includes chimeric antibodies and fully synthetic antibodies, and fragments thereof.

Antibodies are serum proteins whose molecules possess small surface regions that are complementary to chemical groups on their targets. These complementary regions (referred to as antibody binding sites or antigen binding sites), where there are at least 2 complementary regions per antibody molecule and 10, 8 or in some species up to 12 complementary regions per antibody molecule, can react with their corresponding complementary regions (epitopes) on the antigen to join several multivalent antigen molecules together to form a lattice.

The basic building block of a complete antibody molecule consists of 4 polypeptide chains, two identical light (L) chains (each containing about 220 amino acids) and two identical heavy (H) chains (each typically containing about 440 amino acids). The two heavy chains and the two light chains are immobilized by a combination of non-covalent and covalent (disulfide) bonds. The molecule consists of two identical halves, each half having the same antigen binding site consisting of a light chain N-terminal region and a heavy chain N-terminal region. The light and heavy chains typically cooperate to form an antigen-binding surface.

Human antibodies display two types of light chains, κ and λ; a single immunoglobulin molecule is typically only one type or the other. In normal serum, it has been found that 60% of the molecules have kappa determinants and 30% have lambda determinants. Many other species have been found to display both types of light chains, but their proportions vary. For example, in mice and rats, kappa chains account for a small percentage of the total; in dogs and cats, kappa chains are rare; horses do not appear to have any kappa chains; rabbits may have 5 to 40% λ, depending on the strain and b-locus allotype; and chicken light chains are more homologous to λ rather than κ.

In mammals, there are 5 types of antibodies: IgA, IgD, IgE, IgG and IgM, each type having its own class of heavy chains: α (IgA), (IgD), (IgE), γ (IgG) and μ (IgM). In addition, there are 4 IgG immunoglobulin subclasses (IgG1, IgG2, IgG3, IgG4) with γ 1, γ 2, γ 3, and γ 4 heavy chains, respectively. In its secreted form, IgM is a pentamer consisting of 5 four-chain units, giving it a total of 10 antigen binding sites. Each pentamer contains one copy of the J chain, which is covalently inserted between two adjacent tail regions.

All 5 immunoglobulin classes differ from other serum proteins in that they exhibit a wide range of electrophoretic mobilities and are not homogeneous. This heterogeneity, for example individual IgG molecules differ from each other in net charge, is an inherent property of immunoglobulins.

The principle of complementarity, often compared to keys, for fitting in locks involves relatively weak binding forces (hydrophobic and hydrogen bonds, van der waals forces and ionic interactions) that can only function effectively when two reacting molecules are likely to be in close proximity to each other and indeed so close that a protruding constituent atom or group in one molecule can fit into a complementary groove or recess in the other molecule. Antigen-antibody interactions exhibit a high degree of specificity, which is evident at many levels. By specificity, on a molecular level, it is meant that the binding site of an antibody to an antigen has a completely dissimilar complementarity to the antigenic determinants of an unrelated antigen. When antigenic determinants of two different antigens have some structural similarity, it may occur that one determinant somehow fits into the binding site of some antibodies directed against the other antigen, and this phenomenon creates cross-reactivity. Cross-reactivity is important in understanding complementarity or specificity of antigen-antibody reactions. Immunological specificity or complementarity makes it possible to detect small amounts of impurities/contaminants among antigens.

Monoclonal antibodies (mabs) can be produced by: mouse spleen cells derived from immortalized donors are fused with a mouse myeloma cell line to produce stable mouse hybridoma clones that grow in selective media. Hybridoma cells are immortalized hybrid cells resulting from the fusion of antibody-secreting B cells with myeloma cells in vitro. In vitro immortalization, which refers to the initial activation (primary activation) of antigen-specific B cells in culture, is another established method for the production of mouse monoclonal antibodies.

Immunoglobulin heavy chain (V) from peripheral blood lymphocytes_H) And light chain (V)_κAnd V_λ) The diverse library of variable region genes may also be amplified by Polymerase Chain Reaction (PCR) amplification. Genes encoding a single polypeptide chain in which the heavy and light chain variable domains are linked by a polypeptide spacer (single chain Fv or scFv) can be generated by randomly combining the heavy and light chain V genes using PCR. The combinatorial library can then be cloned for display on the surface of a filamentous bacteriophage by fusion with a small capsid protein at the top of the bacteriophage.

The targeted selection technique is based on human immunoglobulin V gene shuffling with rodent immunoglobulin V genes. The method entails (i) shuffling a repertoire of human λ light chains with the heavy chain variable region (VH) domain of a mouse monoclonal antibody reactive with an antigen of interest; (ii) selecting a semi-human Fab on such antigen; (iii) using the selected lambda light chain gene as the "docking domain" of the human heavy chain library in a second shuffling to isolate cloned Fab fragments with human light chain genes; (v) transfecting mouse myeloma cells by electroporation using a mammalian cell expression vector containing the gene; and (vi) expressing the V gene of Fab reactive with this antigen as a whole IgG1, λ antibody molecule in mouse myeloma.

The term "anti-fibrinolytic agent" as used herein refers to a drug used to prevent or dissolve fibrin clots. Antifibrinolytic agents are potent inhibitors of enzymes involved in the fibrinolytic pathway and generally include lysine analogs.

The term "antigen" and its various grammatical forms refer to any substance that can stimulate the production of antibodies and that can specifically bind to these antibodies. The terms "epitope" and "antigenic determinant" are used interchangeably herein to refer to an antigenic site on a molecule that is recognized by an antibody binding site (ACS) and to which the antibody itself binds/attaches. A given epitope may be related to a primary sequence, a secondary sequence, or a tertiary sequence. Sequential antigenic determinants/epitopes are essentially linear chains. In ordered structures such as helical polymers or proteins, an antigenic determinant/epitope is essentially a limited region or patch within or on the surface of the structure involving amino acid side chains from different parts of the molecule that may be in close proximity to each other. They are conformational determinants.

"anti-plasma" is defined as plasma that can be obtained after immunization of a mammal or human with an antigen. It is obtained by separating the particle component from whole blood.

"antisera" is the liquid phase of blood obtained from an immunized mammal (including a human) that is recovered after coagulation has occurred.

"narcotic" refers to a drug that causes a reduction or loss of sensation. Non-limiting examples of anesthetics suitable for use in the context of the present invention include pharmaceutically acceptable salts of lidocaine, bupivacaine, chloroprocaine, dibucaine, etidocaine, mepivacaine, tetracaine, dyclonine, hecaine, procaine, cocaine, ketamine, pramoxine, and phenol.

The term "aneurysm" as used herein means an abnormal enlargement of a blood vessel (artery or vein) in the body, which may occur at any time during the life of the organism.

The term "native" as used herein means derived from the same organism.

The term "biocompatible" as used herein refers to not causing clinically relevant tissue irritation, injury, toxic reactions or immune reactions against living tissue.

The term "biodegradable" as used herein refers to a material that actively or passively decomposes over time by simple chemical treatment, by the action of bodily enzymes, or by other similar bioactive mechanisms.

The term "brain cancer" as used herein refers to metastatic brain cancer that begins at a tumor in the brain or elsewhere in the body and moves to the brain. The term "brain cancer" as used herein includes benign and malignant cancer cells.

The term "carrier" as used herein describes a substance that does not cause significant irritation to an organism and does not impair the biological activity and properties of the compounds in the compositions of the invention. The carriers must be of sufficiently high purity and sufficiently low toxicity to render them suitable for administration to the mammal being treated. The carrier may be inert, or it may possess pharmaceutical benefits. The terms "excipient," "carrier," or "vehicle" are used interchangeably to refer to a carrier substance suitable for formulation and administration of a pharmaceutically acceptable composition as described herein. Carriers and vehicles useful herein include any such materials known in the art to be non-toxic and to interact with other components.

The term "chemotherapeutic agent" in its broadest sense refers to a chemical substance or drug used to treat or control a disease. The term "pool" as used herein means a cavity or enclosed space that acts as a reservoir.

The term "compatible" as used herein means that the components of a composition can be combined with each other in such a way that no interaction exists that would otherwise significantly reduce the efficacy of such composition under ordinary use conditions.

The term "component" as used herein refers to a component part, element or ingredient.

As used herein, the term "condition" relates to a variety of health states, and is intended to include disorders or diseases caused by any underlying mechanism or disorder, damage, and promotion of healthy tissues and organs.

The term "contact" and its full grammatical forms as used herein refers to a state or condition of contact or in close or local proximity.

The term "controlled release" is intended to mean any formulation containing a drug wherein the manner and profile of release of the drug from the formulation is modulated. This refers to immediate release formulations as well as non-immediate release formulations, and non-immediate release formulations include, but are not limited to, sustained release formulations and delayed release formulations.

The term "craniotomy" as used herein refers to the surgical removal of a piece of bone (e.g., a bone fragment) from the skull for surgery on the underlying tissue.

The term "cytokine" as used herein refers to a soluble small protein substance secreted by a cell that has multiple effects on other cells. Cytokines mediate many important physiological functions, including growth, development, wound healing, and immune responses. They act by binding to their cell-specific receptors located in the cell membrane, which allows different signal transduction cascades to be initiated in the cell, which ultimately leads to biochemical and phenotypic changes in the target cell. In general, cytokines act locally. They include type I cytokines, which include numerous interleukins, as well as several hematopoietic growth factors; type II cytokines including interferon and interleukin-10; tumor necrosis factor ("TNF") related molecules, including TNF α and lymphotoxin; immunoglobulin superfamily members, including interleukin 1 ("IL-1"); and chemokines, a family of molecules that play key roles in a variety of immune and inflammatory functions. The same cytokine may exert different effects on a single cell depending on the state of the cell. Cytokines often regulate the expression of other cytokines and trigger cascades of other cytokines.

The term "delayed release" is used herein in its conventional sense to refer to a pharmaceutical formulation wherein there is a time delay between administration of the formulation and release of the drug therefrom. "delayed release" may or may not include gradual release of the drug over an extended period of time, and thus may or may not be "sustained release".

The term "derivative" as used herein means a compound that can be produced in one or more steps from another compound having a similar structure. A "derivative" or "derivatives" of a compound retains at least some degree of the desired function of the compound. Thus, an alternative to a "derivative" may be a "functional derivative".

The term "disease" or "disorder" as used herein refers to an impaired or abnormal functional condition of health.

The term "drug" as used herein refers to a therapeutic agent or any substance other than food that is used in the prevention, diagnosis, alleviation, treatment, or cure of disease.

The term "effective amount" refers to an amount necessary or sufficient to achieve a desired biological effect.

The term "emulsion" as used herein refers to a biphasic system prepared by combining two immiscible liquid carriers, one of which is uniformly dispersed in the other and consists of globules with a diameter equal to or greater than the diameter of the largest colloidal particle. The size of the pellet is critical and must be such that the system achieves maximum stability. Typically, separation of the two phases will occur unless a third material (emulsifier) is incorporated. Thus, the base emulsion contains at least three components, two immiscible liquid carriers and emulsifiers and an active ingredient. Most emulsions incorporate an aqueous phase into a non-aqueous phase (or vice versa). However, substantially anhydrous emulsions can be prepared, for example, anionic and cationic surfactants of the anhydrous immiscible system glycerin and olive oil.

The term "socket" as used herein means a small cavity or depression, as in bone.

The term "hematoma" as used herein refers to a localized mass of extravasated blood that has escaped the vascular constraints present inside surrounding tissue that is relatively or completely constrained inside an organ, tissue, lumen, or potential lumen.

The term "hematoma expansion" as used herein refers to an increase in the volume, size, amount, or range of a hematoma.

The term "hemorrhagic condition" as used herein refers to a disorder or disease in which there is abnormal bleeding.

The term "hydrate" as used herein refers to a compound formed by the addition of water or an element thereof to another molecule. The water can typically be stripped off by heating, thereby producing an anhydrous compound.

The term "hydrogel" as used herein refers to a substance that produces a solid, semi-solid, pseudoplastic, or plastic structure containing the necessary aqueous components to produce a gelatinous or jelly-like substance.

The term "hydrophilic" as used herein refers to a material or substance that has an affinity for a polar substance (e.g., water).

The terms "in vivo", "void volume", "resection pocket", "removal", "injection site", "deposition site", or "implantation site" as used herein are intended to include, without limitation, all tissues of the body, and may refer to any other similar cavity, compartment, or pocket formed by injection, surgical incision, tumor or tissue removal, tissue damage, abscess formation, space formed therein, or thus by clinical assessment of behavior, treatment, or physiological response to a disease or disorder, as non-limiting examples thereof.

The term "injury" as used herein refers to a disruption or damage of a bodily structure or function caused by an external factor or force, which may be physical or chemical injury.

The term "interleukin" as used herein refers to a cytokine secreted by an leukocyte as a means of communicating with other leukocytes.

The term "isolated" is used herein to refer to materials such as, but not limited to, nucleic acids, peptides, polypeptides, or proteins: (1) substantially or essentially free of components that normally accompany or interact with such materials as they exist in their naturally occurring environment. The term "substantially free" or "substantially free" is used herein to mean substantially or significantly free or more than about 95% free or more than about 99% free. The isolated material optionally comprises a material that is not found with the material in nature; or (2) if the substance is in its natural environment, the substance has been synthetically (non-naturally) altered into a composition by deliberate human intervention and/or placed in a location in a cell that is not naturally-occurring relative to the substance present in the environment (e.g., a genomic or subcellular organelle). The alteration that results in such a synthetic substance may be made to a substance that is in or out of its natural state.

The term "isomer" as used herein refers to one of two or more molecules having the same number and type of atoms and thus the same molecular weight, but differing in chemical structure. Isomers may differ in the connectivity of the atoms (structural isomers) or they may have the same connectivity of the atoms, but differ in the arrangement or configuration of the atoms in space (stereoisomers). Stereoisomers may include, but are not limited to, E/Z double bond isomers, enantiomers, and diastereomers. Moieties that may impart stereoisomerism when appropriately substituted include, but are not limited to, olefinic double bonds, imine double bonds or oxime double bonds; tetrahedral carbon, sulfur, nitrogen or phosphorus atoms; and an allene group. Enantiomers are mirror images that cannot be superimposed. A mixture of equal parts of the optical form of one compound is called a racemic mixture or racemate. Diastereomers are stereoisomers that are not mirror images. The present invention provides each pure stereoisomer of any compound described herein. Such stereoisomers may include enantiomers, diastereomers, or E or Z alkene, imine or oxime isomers. The invention also provides stereoisomeric mixtures, including racemic, diastereomeric, or E/Z isomeric mixtures. Stereoisomers may be synthesised in pure form (Nmogr' di, M.; Stereoselective Synthesis, (1987) VCH, editors Ebel, H. and Asymmetric Synthesis, Vol.3-5, (1983) Academic Press, editors Morrison, J.), or they may be resolved by a variety of methods, such as crystallisation and chromatographic techniques (Jaques, J.; Collet, A.; Wilen, S.; Enantiomer, peptides, and solutions,1981, John Wiley and Sons and Asymmetric Synthesis, Vol.2, 1983, Academic Press, edison, J.). Furthermore, the compounds of the invention may exist as enantiomers, diastereomers, isomers, or two or more of the compounds may exist as racemic or diastereomeric mixtures.

The term "labile" as used herein means susceptible to increased degradation.

The term "lipophilic" as used herein means that a non-polar environment is favored or possesses an affinity for a non-polar environment as compared to a polar or aqueous environment.

As used herein, the term "long-term" release refers to the construction and placement of an implant to deliver therapeutic levels of an active ingredient for at least 7 days and possibly up to about 30 days to about 60 days.

The term "minimizing progression" as used herein refers to reducing the amount, extent, size, or degree of progression of a group or series of events.

The term "adjust" as used herein refers to adjust, change, modify or adjust to a certain amount or proportion. This modulation can be any change, including an undetectable change.

The term "parenteral" as used herein refers to introduction into the body by injection (i.e., administration by injection), including, for example, subcutaneous (i.e., injection under the skin), intramuscular (i.e., injection into muscle); intravenous (i.e., injection into a vein), intrathecal (i.e., injection into the space around the spinal cord or under the arachnoid membranes of the brain), intrasternal injection, or infusion techniques. Compositions for parenteral administration are delivered using a needle (e.g., a surgical needle). The term "surgical needle" as used herein refers to any needle adapted to deliver a fluid (i.e., flowable) composition to a selected anatomical structure. Injectable preparations, for example sterile injectable aqueous or oleaginous suspensions, may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents.

The term "particle" as used herein refers to a very small component, e.g., a nanoparticle or microparticle, that may contain, in whole or in part, at least one therapeutic agent as described herein.

The term "improving patient dosing outcome" as used herein refers to the absence or reduction of at least one side effect accompanying systemic administration of an anti-fibrinolytic agent. Examples of side effects include, but are not limited to, hypertension, cardiac arrhythmias, edema, rhabdomyolysis, thrombosis, cerebral infarction or stroke, and myocardial infarction or heart attack.

The term "pharmaceutical composition" is used herein to refer to a composition that is used to prevent, reduce the strength of, cure, or otherwise treat a target condition or disease.

The term "pharmaceutically acceptable carrier" as used herein refers to any substantially non-toxic carrier in which the product of the invention will remain stable and biologically effective that can be used to formulate and administer the compositions of the invention. The pharmaceutically acceptable carrier must be of sufficiently high purity and sufficiently low toxicity to render it suitable for administration to the mammal being treated. It should also maintain stability and bioavailability of the active agent. When combined with the active agent and other components of a given composition, the pharmaceutically acceptable carrier can be liquid or solid and is selected to provide the desired volume, consistency, etc. according to the intended mode of administration in mind. The term "pharmaceutically acceptable salts" means those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.

The term "periosteum" as used herein refers to the normal enclosure of bone, consisting of a dense fibrous outer layer to which muscle is attached and a more fragile inner layer capable of forming bone.

The term "preventing" as used herein means to retain, hinder or prevent an event, action or action from occurring, occurring or emerging.

The term "prodrug" as used herein means a peptide or derivative that is in an inactive form and is converted to an active form by biotransformation after administration to a subject.

The term "solvate" as used herein refers to a complex formed by the attachment of solvent molecules to a solute. The term "solvent" refers to a substance that is capable of dissolving another substance (referred to as a "solute") to form a uniformly dispersed mixture (solute).

The term "recombinant" as used herein refers to a substance produced by genetic engineering.

The term "reduce" or "to reduce" as used herein refers to a reduction, attenuation, or attenuation in degree, intensity, range, size, amount, density, or number.

The term "similar" is used interchangeably with the terms "similar", "comparable" or "like", and means having a common trait or characteristic.

The term "sinus" and its various grammatical forms refer to a conduit (canal) or a dilated area in a tube.

The terms "soluble" and "solubility" refer to the property of being readily soluble in a given fluid (solvent). The term "insoluble" refers to the property of a substance that has minimal or limited solubility in a given solvent. In solution, the molecules of the solute (or dissolved substance) are uniformly distributed among the molecules of the solvent. A "suspension" is a dispersion (mixture) in which a finely divided species is combined with another species, the former being so finely dispersed and mixed that it does not settle rapidly. In daily life, the most common suspensions are those of solids in liquids. Among the acceptable vehicles and solvents that may be employed are water, ringer's solution and isotonic sodium chloride solution.

The term "susceptible" as used herein refers to a member of a population at risk.

The terms "subject" or "individual" or "patient" are used interchangeably to refer to a member of a mammalian species, including humans.

The phrase "subject with a subdural hematoma" refers to a subject who exhibits diagnostic markers and symptoms associated with SDH.

The term "sustained release" (also referred to as "extended release") is used herein in its conventional sense to refer to a pharmaceutical formulation that provides gradual release of the drug over an extended period of time and preferably, although not necessarily, produces substantially constant blood levels of the drug over an extended period of time. Alternatively, delayed absorption of a parenterally administered drug form is achieved by dissolving or suspending the drug in an oil vehicle. Non-limiting examples of sustained release biodegradable polymers include polyesters, polyester polyethylene glycol copolymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, SAIBs, photopolymerizable biopolymers, protein polymers, collagen, polysaccharides, chitosan, and alginates.

The term "symptom" as used herein refers to a phenomenon attributed to or accompanied by a particular disease or disorder and serving as an indication thereof.

The term "syndrome" as used herein refers to a pattern of symptoms that indicate a certain disease or condition.

The term "therapeutic agent" as used herein refers to a drug, molecule, nucleic acid, protein, composition, or other substance that provides a therapeutic effect. The term "active" as used herein refers to an ingredient, component, or constituent of a composition of the present invention that is responsible for the intended therapeutic effect. The terms "therapeutic agent" and "active agent" are used interchangeably. The term "therapeutic component" as used herein refers to a therapeutically effective dose (i.e., dose and frequency of administration) that eliminates, reduces, or prevents progression of a particular disease manifestation, at a certain percentage of the population. An example of a commonly used therapeutic component is ED₅₀It describes an amount that appears therapeutically effective in a particular dose for a particular disease in 50% of the population.

The term "therapeutic effect" as used herein refers to the outcome of a treatment that is judged to be desirable and beneficial. Therapeutic effects may include directly or indirectly stasis, reduction or elimination of disease manifestations. Therapeutic effects may also include directly or indirectly arresting, reducing or eliminating the progression of disease manifestations.

The term "therapeutically effective amount" or "effective amount" of one or more active agents is an amount sufficient to provide the desired therapeutic benefit. An effective amount of active agent that can be used is generally from about 0.1mg/kg body weight to about 50mg/kg body weight. However, the dosage level will depend upon a variety of factors including the type of injury, age, body weight, sex, medical condition of the patient, the severity of the condition, the route of administration and the particular active agent used. Thus, the dosage regimen may vary widely, but may be routinely determined by the surgeon using standard methods.

The term "thickener" as used herein refers to a substance that makes the compositions of the present invention dense or viscous in consistency.

The term "transgenic" as used herein refers to an experimental strain of an animal that has been genetically modified in the laboratory to add or delete a particular exogenous gene to the DNA of the animal. Common transgenic animal models include, but are not limited to, mice.

Traumatic Brain Injury (TBI) results from head injury that can cause persistent damage to the brain and affect up to ten million patients worldwide each year. The health effects of TBI can be debilitating, leading to long-term disability and creating a significant financial burden.

Traumatic brain injury is caused by external mechanical forces (e.g., head strike, impact force, acceleration-deceleration force, or slinging). It can occur when the skull is fractured and the brain is punctured directly (open head injury) and also when the skull remains intact but the brain is still subject to damage (closed head injury).

Symptoms of TBI vary in severity depending on the extent of the injured brain, and may include headache, neck pain, confusion, memory difficulties, difficulty concentrating or deciding, dizziness, weakness, mood changes, nausea, irritability, photophobia, blurred vision, tinnitus, loss of taste or smell, seizures (seizure), sleep disorders, hypoxemia, hypotension, and brain swelling.

TBI is rated as mild (meaning a brief change in mental state or consciousness), moderate or severe (meaning a prolonged period of unconsciousness or amnesia after injury) based on the level of consciousness after resuscitation or Glasgow Coma Scale (GCS) score. GCS assesses open eyes (spontaneous 4, speech 3, pain 3, no 1), motor responses (obedience 6, localization 5, withdrawal 4, abnormal depression 3, extensor response 2, no 1) and verbal responses (directional 5, confusion 4, unsuitability 3, fiddle 2, no 1). Mild TBI (GCS 13-15) concussions in most cases and there is complete neurological recovery, although many of these patients have short-term memory difficulties and concentration difficulties. In moderate TBI (GCS 9-13), the patient sleeps or is stiff, and in severe injury (GCS 3-8), the patient is unconscious, unable to open his or her eyes, or is subject to commands.

Patients with severe TBI (lethargy) have significant risk of hypotension, hypoxemia and brain swelling. If these sequelae are not properly prevented or treated, they can exacerbate brain damage and increase the risk of death.

The term "traumatic intracerebral hemorrhage" (ICH) as used herein refers to such hemorrhage caused by or associated with traumatic injury.

The terms "treat" or "treating" include eliminating, substantially inhibiting, delaying or reversing the progression of a disease, condition, or disorder, substantially ameliorating the clinical or aesthetic symptoms of a condition, substantially preventing the appearance of the clinical or aesthetic symptoms of a disease, condition, or disorder, and protecting from harmful or unpleasant symptoms. The term "treating" or "treating" as used herein further refers to achieving one or more of the following: (a) reducing the severity of the condition; (b) limiting the development of symptoms characteristic of the disorder being treated; (c) limiting the worsening of symptoms characteristic of the condition being treated; (d) limiting recurrence of the disorder in a patient who has had the disorder; and (e) limiting recurrence of symptoms in patients who previously had symptoms of the disorder.

The term "topical" refers to the application of a composition at or immediately below the point of application. The phrase "topically applying" describes applying to one or more surfaces, including epithelial surfaces. However, in contrast to transdermal administration, topical administration generally provides a local effect rather than a systemic effect. As used herein, the terms "topical administration" and "transdermal administration" are used interchangeably unless otherwise indicated or suggested.

The term "whole blood" as used herein refers to generally raw or unmodified collected blood containing all components, including, but not limited to, plasma, cellular components (e.g., erythrocytes, leukocytes (including lymphocytes, monocytes, eosinophils, basophils, and neutrophils), and platelets), proteins (e.g., fibrinogen, albumin, immunoglobulins), hormones, clotting factors, and fibrin factors. The term "whole blood" includes any anticoagulant that may be combined with blood at the time of blood collection.

The term "vascular malformation" as used herein refers to a malformation of vascular development in the brain that results in abnormal accumulation or patterning of blood vessels.

The term "xenogeneic" as used herein refers to belonging to different species.

I. Non-human animal model of hemorrhagic encephalopathy

According to one aspect, the invention provides a non-human animal model system for hemorrhagic brain disease. According to some such embodiments, the hemorrhagic brain condition is chronic SDH. According to some such embodiments, the hemorrhagic brain condition is ICH. According to one embodiment, the non-human animal model system provides for administering to a mammal an initiator composition that induces a hemorrhagic brain condition.

According to another aspect, the invention provides a mammal having an induced hemorrhagic encephalopathy, resulting in a hematoma. According to some such embodiments, the hemorrhagic brain condition is chronic SDH. According to some such embodiments, the hemorrhagic brain condition is ICH. According to some embodiments, the hematoma remains stable over time. According to some embodiments, the hematoma expands over time. According to some embodiments, the expansion of the hematoma is progressive.

According to some embodiments, the mammal is a mouse. According to another embodiment, the mammal is a transgenic mouse. According to another embodiment, the mammal is a rat. According to another embodiment, the mammal is a member of the order Rodentia (Rodentia).

According to another embodiment, the initiator composition is administered into the subcutaneous cavity of the mammal.

According to another embodiment, the initiator composition is administered intracranially. According to some embodiments, the initiator composition is administered proximate to the dura mater of the mammal. According to some embodiments, the initiator composition is administered by surgical injection. According to some embodiments, the initiator composition is deposited on or within the implant.

According to another embodiment, the initiator composition comprises a fluid obtained from a chronic SDH. According to another embodiment, the initiator composition comprises autologous blood or a component thereof. According to another embodiment, the initiator composition comprises allogeneic blood or a component thereof. According to another embodiment, the initiator composition comprises xenogeneic blood or a component thereof. According to another embodiment, the initiator composition comprises an antibody directed against at least one coagulation factor. According to another embodiment, the initiator composition comprises an enzyme that catalyzes the breakdown of collagen, such as collagenase. According to another embodiment, the initiator composition comprises antibodies against at least one coagulation factor selected from the group consisting of procoagulants, anticoagulants, clot structural factors, fibrin factors, and phospholipids. According to another embodiment, the procoagulant is selected from the group consisting of coagulation factor II, factor V, factor VII, factor IX, factor X, factor XI, factor XII, prekallikrein, kininogen and tissue factor. According to another embodiment, the anticoagulant is selected from the group consisting of protein C, protein S, antithrombin III and heparin cofactor II. According to another embodiment, the clot structural factor is selected from fibrinogen and factor XIII. According to another embodiment, the fibrinolytic factor is selected from the group consisting of plasminogen, tissue-type plasminogen activator, plasminogen activator inhibitor and alpha 2-plasmin inhibitor. According to another embodiment, the initiator composition for inducing chronic SDH in a non-human animal model comprises whole blood. According to some embodiments, the whole blood is autologous. According to some embodiments, the whole blood is allogeneic. According to some embodiments, the whole blood is xenogeneic. According to some such embodiments, the blood has a volume of 1 μ Ι to about 20 ml. According to some such embodiments, the blood has a volume of 100 μ Ι to about 15 ml. According to some such embodiments, the blood has a volume of 500 μ Ι to about 12.5 ml. According to some such embodiments, the blood has a volume of 1ml to about 10 ml. According to some such embodiments, the volume of blood is about 2 ml. According to some such embodiments, the volume of blood is about 3 ml. According to some such embodiments, the volume of blood is about 4 ml. According to some such embodiments, the volume of blood is about 5 ml. According to some such embodiments, the volume of blood is about 6 ml. According to some such embodiments, the volume of blood is about 7 ml. According to some such embodiments, the volume of blood is about 8 ml. According to some such embodiments, the volume of blood is about 9 ml. According to some such embodiments, the initiator composition comprises at least one component of whole blood. Examples of components of whole blood include, but are not limited to, plasma, cellular components (e.g., erythrocytes, leukocytes (including lymphocytes, monocytes, eosinophils, basophils, and neutrophils), and platelets), proteins (e.g., fibrinogen, albumin, immunoglobulins), hormones, clotting factors, and fibrin factors.

According to another embodiment, the initiator composition for inducing haematoma-causing hemorrhagic encephalopathy in a non-human animal model comprises a fluid obtained from chronic SDH. According to some such embodiments, the fluid obtained from the chronic SDH is in a volume of about 1 μ l to about 20 ml. According to some such embodiments, the fluid obtained from the chronic SDH is in a volume of about 100 μ l to about 15 ml. According to some such embodiments, the fluid obtained from the chronic SDH is in a volume of about 500 μ l to about 12.5 ml. According to some such embodiments, the fluid obtained from the chronic SDH is in a volume of about 1ml to about 10 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 2 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 3 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 4 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 5 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 6 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 7 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 8 ml. According to some such embodiments, the volume of fluid obtained from a chronic SDH is about 9 ml.

According to another embodiment, the initiator composition for inducing a hemorrhagic encephalopathy causing a hematoma in a non-human animal model comprises an antibody.

According to another embodiment, the initiator composition for inducing a hemorrhagic encephalopathy causing hematoma in a non-human animal model is a monoclonal antibody. According to another embodiment, the initiator for inducing a hemorrhagic encephalopathy causing a hematoma in a non-human animal model is a polyclonal antibody.

According to another embodiment, the initiator composition for inducing haematoma-causing hemorrhagic encephalopathy in a non-human animal model comprises a monospecific antibody directed against one coagulation factor or against a specific epitope of such a coagulation factor. According to another embodiment, the initiator composition for inducing a hemorrhagic brain condition causing a hematoma in a non-human animal model comprises an antibody recognizing at least two coagulation factors. For example, the initiator composition may comprise an antibody to human plasmin or human plasminogen that cross-reacts with animal-derived plasmin or plasminogen. Alternatively, the initiator composition may comprise an antibody against human thrombin that is cross-reactive with animal thrombin. Alternatively, the initiator composition may comprise antibodies against vitamin K-dependent blood factors (e.g., factors of the prothrombin complex).

According to other embodiments, the initiator composition comprises antibodies against at least one procoagulant (including, but not limited to, coagulation factors II, V, VII, IX, X, XI, XII, prekallikrein, kininogen, and tissue factor), anti-coagulants (including, but not limited to, protein C, protein S, antithrombin III, heparin cofactor II), coagulation structural factors (including, but not limited to, fibrinogen and factor XIII), fibrinolytic factors (such as, but not limited to, plasminogen, t-PA, PAI-1, and α 2-plasmin inhibitors), and phospholipids.

The suitability of an antibody preparation for this purpose can be determined based on a variety of assays. For example, an in vitro assay may be performed in which an antibody preparation is incubated with a cerebrospinal fluid sample of a test animal and inhibition or elimination of a blood factor is determined.

The desired effect of altered chronic SDH recurrence can be shown in vivo in test animals by measuring hematoma, its rate of formation or dissolution.

According to some embodiments, the initiator composition may be prepared by: immunizing a mammal with plasma, a plasma fraction or a recombinant equivalent thereof, recovering the anti-plasma or anti-serum and subsequently absorbing one or several antibodies of such anti-plasma or anti-serum, whereby said initiator composition will only contain such functional antibodies that can selectively functionally inhibit and/or eliminate at least one blood factor in the mammal.

The process for preparing such initiator composition comprises the steps of: (a) immunizing a mammal with plasma, a plasma fraction or a recombinant equivalent thereof, (b) recovering an anti-plasma or anti-serum from the immunized animal of (a), c) optionally purifying the antibody fraction from the anti-plasma or anti-serum of (a), and (d) formulating a composition suitable for infusion into or near the dura mater, brain or subcutaneous tissue of the mammal.

According to some embodiments, the initiator composition may be prepared by: immunizing a mammal with fluid obtained from a human chronic SDH, a fraction of fluid obtained from a human chronic SDH or a recombinant equivalent thereof, recovering the anti-plasma or antiserum and subsequently purifying the antibodies from such anti-plasma or antiserum, whereby said initiator composition will contain only those functional antibodies that can selectively inhibit and/or eliminate at least one blood factor in the mammal.

The process for preparing such initiator composition comprises the steps of: (a) immunizing a mammal with human chronic SDH fluid, a fraction of human chronic SDH fluid or a recombinant equivalent thereof, (b) recovering the anti-plasma or anti-serum from the immunized animal of (a), c) optionally purifying the anti-plasma or anti-serum antibody fraction of (a), and (d) formulating a composition suitable for infusion into or near the dura mater of a mammal.

According to another embodiment, chronic SDH or similar disorders can be more precisely caused by preparing an initiator composition with antibodies specific for a certain blood factor, wherein said antibodies do not cross-react with other blood factors that may be contained in the immunological material. Mammals that can be immunized include, but are not limited to, sheep, goats, cattle, pigs, rabbits, guinea pigs, horses, rats, and mice. Mammals in which chronic SDH can be induced include, but are not limited to, mice, rats, and other members of the rodent order.

According to another embodiment, the invention provides a method of treating a mammal that is a model for blood factor deficiency by anti-plasma antibodies (e.g., in an infusion composition) by: functionally inhibiting and/or eliminating several blood factors, thereby altering the extracorporeal or ex vivo clotting time or the recurrence of chronic SDH in such mammals. According to some embodiments, antibodies to certain blood factors are adsorbed in vivo by administering the blood factors to an animal.

According to some embodiments, blood factors can be reconstructed and altered clotting times and characteristics of chronic SDH can be identified as being dependent on the reconstructed blood factors.

According to another embodiment, the invention further provides a method for determining the characteristics of recurrent chronic SDH, the method comprising the steps of: (a) inducing chronic SDH in mammals; (b) collecting fluid obtained from the chronic SDH; (c) determining the blood factor component of the fluid of step (b); and (d) determining the rate at which the size of the chronic SDH expands or contracts over time.

According to another embodiment, the initiator composition further comprises a carrier. According to another embodiment, the carrier is a pharmaceutically acceptable carrier. According to some embodiments, the initiator is in the form of a pharmaceutical composition.

According to another embodiment, the initiator composition is in an amount of about 0.000001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000003mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0003mg to about 10g per kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.001mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.01mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 10mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 20mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 30mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 40mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 50mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 60mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 70mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 80mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 90mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 100mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 110mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 120mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 130mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 140mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 150mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 160mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 170mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 180mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 190mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 200mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 250mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 500mg/kg body weight.

Methods for evaluating substances for treating a hemorrhagic condition in the brain

According to another aspect, the invention provides a method for evaluating the ability of a substance to treat recurrent bleeding in a mammal due to a hemorrhagic condition of the brain, the method comprising the steps of: (a) administering a substance to be evaluated to a mammal; and (b) measuring a change in at least one coagulation parameter, at least one fibrinolysis parameter (meaning one of a set of measurable factors) or at least one parameter characteristic of a hemorrhagic condition in the mammal by the substance to be assessed. The coagulation parameter or fibrinolysis parameter may be determined according to known methods. For example, a series of in vitro tests for testing blood coagulation factors or fibrinolytic factors and their role in samples are known.

According to some embodiments, the mammal is a mouse. According to another embodiment, the mammal is a knockout mouse. According to another embodiment, the mammal is a rat. According to another embodiment, the mammal is a member of the order rodentia.

According to one embodiment, the hemorrhagic condition of the brain is chronic SDH. According to one embodiment, the method further comprises the step of treating the mammal with an initiator composition for inducing chronic SDH. According to another embodiment, the initiator composition for inducing chronic SDH in a non-human animal model comprises whole blood. According to some embodiments, the whole blood is autologous. According to some embodiments, the whole blood is allogeneic. According to some embodiments, the whole blood is xenogeneic. According to some such embodiments, the blood is in a volume of about 1 μ l to about 20 ml. According to some such embodiments, the blood is in a volume of about 100 μ l to about 15 ml. According to some such embodiments, the blood is in a volume of about 500 μ Ι to about 12.5 ml. According to some such embodiments, the blood is in a volume of about 1ml to about 10 ml. According to some such embodiments, the volume of blood is about 2 ml. According to some such embodiments, the volume of blood is about 3 ml. According to some such embodiments, the volume of blood is about 4 ml. According to some such embodiments, the volume of blood is about 5 ml. According to some such embodiments, the volume of blood is about 6 ml. According to some such embodiments, the volume of blood is about 7 ml. According to some such embodiments, the volume of blood is about 8 ml. According to some such embodiments, the volume of blood is about 9 ml. According to some such embodiments, the initiator composition comprises at least one component of whole blood. Examples of components of whole blood include, but are not limited to, plasma, cellular components (e.g., erythrocytes, leukocytes (including lymphocytes, monocytes, eosinophils, basophils, and neutrophils), and platelets), proteins (e.g., fibrinogen, albumin, immunoglobulins), hormones, clotting factors, and fibrin factors.

According to another embodiment, the step (a) of applying the substance to be evaluated to the mammal comprises applying the initiator composition to a rodent. According to some such embodiments, the initiator composition comprises 6-aminonicotinamide. According to some such embodiments, the dose of 6-aminonicotinamide is 0-10mg/kg body weight. According to some such embodiments, the dose of 6-aminonicotinamide is 11-20mg/kg body weight. According to some such embodiments, the dose of 6-aminonicotinamide is 21-30mg/kg body weight.

According to another embodiment, the initiator composition for inducing chronic SDH comprises a fluid obtained from human chronic SDH. According to some such embodiments, the fluid obtained from human chronic SDH is in a volume of about 1 μ l to about 20 ml. According to some such embodiments, the fluid obtained from human chronic SDH is in a volume of about 100 μ l to about 15 ml. According to some such embodiments, the fluid obtained from human chronic SDH is in a volume of about 500 μ l to about 12.5 ml. According to some such embodiments, the fluid obtained from human chronic SDH is in a volume of about 1ml to about 10 ml. According to some such embodiments, the fluid obtained from human chronic SDH is about 2 ml. According to some such embodiments, the fluid obtained from human chronic SDH is about 3 ml. According to some such embodiments, the fluid obtained from human chronic SDH is about 4 ml. According to some such embodiments, the fluid obtained from human chronic SDH is about 5 ml. According to some such embodiments, the volume of cerebrospinal fluid is about 6 ml. According to some such embodiments, the volume of cerebrospinal fluid is about 7 ml. According to some such embodiments, the volume of cerebrospinal fluid is about 8 ml. According to some such embodiments, the volume of cerebrospinal fluid is about 9 ml.

According to some embodiments, the initiator composition is in the form of a pharmaceutical composition. According to some embodiments, the initiator composition further comprises a carrier. According to some embodiments, the carrier is a pharmaceutically acceptable carrier.

According to another embodiment, the initiator composition used to induce chronic SDH is an antibody. According to another embodiment, the initiator composition used to induce chronic SDH is a monoclonal antibody. According to another embodiment, the initiator composition used to induce chronic SDH is a polyclonal antibody. According to some embodiments, the initiator composition is an anti-plasma antibody preparation.

According to some embodiments, the initiator composition comprises an antibody directed against at least one procoagulant (including, but not limited to, coagulation factors II, V, VII, IX, X, XI, XII, prekallikrein, kininogen, and tissue factor), an antibody directed against an anticoagulant (including, but not limited to, protein C, protein S, antithrombin III, heparin cofactor II), a coagulation structural factor (including, but not limited to, fibrinogen and factor XIII), a fibrinolytic factor (such as, but not limited to, plasminogen, tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor (PAI-1), and α 2-plasmin inhibitor), and a phospholipid.

According to some embodiments, the magnitude of at least one parameter characteristic of chronic SDH of a mammal suffering from inducible chronic SDH is compared with the magnitude of at least one parameter characteristic of chronic SDH of a mammal to which a compound or substance to be tested has been administered, in order to determine the extent to which the substance to be tested can treat inducible chronic SDH. Examples of such parameters include, but are not limited to, bleeding behavior, blood volume in chronic SDH, size of hematoma area, expansion or contraction of hematoma area, expansion kinetics of chronic SDH, and contraction kinetics of chronic SDH. According to some embodiments, the compound or substance to be tested is administered before the induction of chronic SDH. According to some embodiments, the compound or substance to be tested is applied simultaneously with the initiator composition. According to some embodiments, the compound or substance to be tested is administered after the induction of chronic SDH.

According to another embodiment, the initiator composition is applied to the subcutaneous cavity of the dorsal surface of the mammal. According to another embodiment, the initiator composition is administered to the brain of the mammal. According to some such embodiments, the initiator composition is administered to the brain after a craniotomy. According to some such embodiments, the initiator composition is administered to the brain through a borehole. The term "craniotomy" as used herein refers to any bony opening (bony opening) cut into the skull. A piece of skull bone (called a bone fragment) is removed to reach the brain below. There are many types of craniotomies, which are named according to the region of the skull to be removed. According to some such embodiments, the bore is a brow bore. According to some such embodiments, the bore is a parietal bore. According to some such embodiments, the bore is a temporal bore. According to some such embodiments, the bore is an occipital bore. Some common craniotomies include frontotemporal craniotomies, parietal craniotomies, temporal craniotomies, and subcapillary craniotomies. Typically, the bone fragments are replaced. According to some embodiments, stereotactic frames, video-guided computer systems, or endoscopes are used to precisely guide instruments through a borehole.

No food and water is allowed for 8 hours prior to surgery. General anesthesia will be administered. Once asleep, the animal's head will be placed in a skull fixation device to secure the head in place during surgery.

After the hair is shaved off of the intended incision area and the scalpel is primed with a preservative, a skin incision will be made. The skin and muscles lift away from the bone and fold back. Next, a surgical drill will be used to create one or more small burr holes in the skull. In some embodiments, one bone fragment will be produced. The cut piece of bone will be lifted and removed to expose the dura mater. The bone fragment will be safely stored until it is replaced at the end of the surgery. In some cases, the drain may be placed under the skin for several days to remove blood or fluid from the surgical field. The muscle and skin will be sutured back together and a soft adhesive dressing will be placed over the incision.

According to some embodiments, the initiator composition is administered intracranially. According to some such embodiments, the initiator composition is administered by perfusion. According to some such embodiments, the initiator composition is administered subdurally. According to some such embodiments, the initiator composition is administered intracerebrally. According to some such embodiments, the initiator composition is administered by surgical injection. According to some such embodiments, the initiator composition is deposited on or within the implant.

According to another embodiment, the initiator composition is in an amount of about 0.000001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000003mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0003mg to about 10g per kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.001mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.01mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 10mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 20mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 30mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 40mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 50mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 60mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 70mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 80mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 90mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 100mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 110mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 120mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 130mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 140mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 150mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 160mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 170mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 180mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 190mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 200mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 250mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 500mg/kg body weight.

According to another embodiment, the hemorrhagic condition of the brain is an intracerebral hematoma. According to one embodiment, the method further comprises the step of treating the mammal with an initiator composition for inducing intracerebral hematoma. According to another embodiment, the initiator composition for inducing intracerebral hematoma in a non-human animal model comprises whole blood. According to some embodiments, the whole blood is autologous. According to some embodiments, the whole blood is allogeneic. According to some embodiments, the whole blood is xenogeneic. According to some such embodiments, the blood is in a volume of about 1 μ l to about 20 ml. According to some such embodiments, the blood is in a volume of about 100 μ l to about 15 ml. According to some such embodiments, the blood is in a volume of about 500 μ Ι to about 12.5 ml. According to some such embodiments, the blood is in a volume of about 1ml to about 10 ml. According to some such embodiments, the volume of blood is about 2 ml. According to some such embodiments, the volume of blood is about 3 ml. According to some such embodiments, the volume of blood is about 4 ml. According to some such embodiments, the volume of blood is about 5 ml. According to some such embodiments, the volume of blood is about 6 ml. According to some such embodiments, the volume of blood is about 7 ml. According to some such embodiments, the volume of blood is about 8 ml. According to some such embodiments, the volume of blood is about 9 ml. According to some such embodiments, the initiator composition comprises at least one component of whole blood. Examples of components of whole blood include, but are not limited to, plasma, cellular components (e.g., erythrocytes, leukocytes (including lymphocytes, monocytes, eosinophils, basophils, and neutrophils), and platelets), proteins (e.g., fibrinogen, albumin, immunoglobulins), hormones, clotting factors, and fibrin factors.

According to another embodiment, the initiator composition for inducing intracerebral hematoma comprises collagenase. As used herein, one unit (U) of collagen activity dissolves 1mg of collagen fibrils per hour at 37 ℃. According to some embodiments, the initiator composition comprises collagenase in an amount from about 0.001U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 0.01U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 0.1U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 1U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 25U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 50U/mg body weight to about 100U/mg body weight. According to some embodiments, the initiator composition comprises collagenase in an amount from about 75U/mg body weight to about 100U/mg body weight.

According to another embodiment, the initiator composition for inducing intracerebral hematoma comprises at least one antibody. According to another embodiment, the initiator composition for inducing intracerebral hematoma comprises at least one monoclonal antibody. According to another embodiment, the initiator composition for inducing intracerebral hematoma comprises at least one polyclonal antibody. According to some embodiments, the initiator composition comprises an anti-plasma antibody preparation.

According to other embodiments, the initiator composition comprises antibodies against at least one procoagulant (including, but not limited to, coagulation factors II, V, VII, IX, X, XI, XII, prekallikrein, kininogen, and tissue factor), anti-coagulants (including, but not limited to, protein C, protein S, antithrombin III, heparin cofactor II), coagulation structural factors (including, but not limited to, fibrinogen and factor XIII), fibrinolytic factors (such as, but not limited to, plasminogen, t-PA, PAI-1, and α 2-plasmin inhibitors), and phospholipids.

According to some embodiments, at least one parameter characteristic of an intracerebral hematoma in a mammal having an induced intracerebral hematoma is compared to at least one parameter characteristic of an intracerebral hematoma in a mammal to which a test compound or substance has been administered in order to determine the extent to which the test substance can treat an induced intracerebral hematoma. Examples of parameters that are characteristic of an intracerebral hematoma include, but are not limited to, bleeding behavior, blood volume in an intracerebral hematoma, size of a hematoma region, expansion or contraction of a hematoma region, expansion kinetics of an intracerebral hematoma, and contraction kinetics of an intracerebral hematoma. According to some embodiments, the agent is administered prior to inducing an intracerebral hematoma. According to some embodiments, the substance is applied simultaneously with the initiator composition. According to some embodiments, the agent is administered after induction of an intracerebral hematoma.

According to another embodiment, the initiator composition is administered to the brain of the mammal. According to some such embodiments, the initiator composition is administered to the brain after a craniotomy. According to some such embodiments, the initiator composition is administered to the brain through a borehole. According to some such embodiments, the bore is a brow bore. According to some such embodiments, the bore is a parietal bore. According to some such embodiments, the bore is a temporal bore. According to some such embodiments, the bore is an occipital bore. According to some embodiments, stereotactic frames, video-guided computer systems, or endoscopes are used to precisely guide instruments through a borehole.

No food and water is allowed for 8 hours prior to surgery. General anesthesia will be administered. Once asleep, the animal's head will be placed in a skull fixation device to secure the head in place during surgery.

After the hair is shaved off of the intended incision area and the scalpel is primed with a preservative, a skin incision will be made. The skin and muscles lift away from the bone and fold back. Next, a surgical drill will be used to create one or more small burr holes in the skull. In some embodiments, one bone fragment will be produced. The cut piece of bone will be lifted and removed to expose the dura mater. The bone fragment will be safely stored until it is replaced at the end of the surgery. In some cases, the drain may be placed under the skin for several days to remove blood or fluid from the surgical field. The muscle and skin will be sutured back together and a soft adhesive dressing will be placed over the incision.

According to some embodiments, the initiator composition is administered intracranially. According to some such embodiments, the initiator composition is administered by perfusion. According to some such embodiments, the initiator composition is administered subdurally. According to some such embodiments, the initiator composition is administered by surgical injection. According to some such embodiments, the initiator composition is deposited on or within the implant.

According to another embodiment, the initiator composition is in an amount of about 0.000001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000003mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.000009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0003mg to about 10g per kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.00009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.0005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.001mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.005mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.01mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 0.1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 1mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 10mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 20mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 30mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 40mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 50mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 60mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 70mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 80mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 90mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 100mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 110mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 120mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 130mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 140mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 150mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 160mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 170mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 180mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 190mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 200mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 250mg/kg body weight. According to another embodiment, the initiator composition is in an amount of about 500mg/kg body weight.

Pharmaceutical composition for treating hemorrhagic conditions of the brain

According to another aspect, the invention provides a pharmaceutical composition for treating a hemorrhagic condition of the brain in a mammal, the pharmaceutical composition comprising a therapeutically effective amount of a therapeutic agent and a pharmaceutically acceptable carrier. According to one embodiment, the hemorrhagic condition of the brain is chronic SDH. According to one such regimen, the pharmaceutical composition is a pharmaceutical composition for administration into or near the dura mater. According to another embodiment, the hemorrhagic condition of the brain is ICH. According to one such regimen, the pharmaceutical composition is a pharmaceutical composition for administration into or near a site of ICH.

According to one embodiment, the therapeutic agent comprises an antifibrinolytic agent or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises aminocaproic acid, a functional derivative of aminocaproic acid, or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises factor VII. According to another embodiment, the therapeutic agent comprises recombinant factor VII. According to another embodiment, the therapeutic agent comprises tranexamic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises aprotinin. According to another embodiment, the therapeutic agent comprises antiplasmin or an ester, salt, hydrate, solvate, functional derivative, or prodrug thereof. According to another embodiment, the therapeutic agent comprises fibrin fragment D or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises vitamin K or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises vitamin K₁Or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises vitamin K₂Or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises vitamin K₃Or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to another embodiment, the therapeutic agent comprises 4-aminomethylbenzoic acid or an ester, salt, hydrate, solvent thereofA compound, a functional derivative or a prodrug.

According to another embodiment, the pharmaceutically acceptable carrier is a gel compound.

According to another embodiment, the pharmaceutically acceptable carrier is a semi-solid compound.

According to another embodiment, the pharmaceutically acceptable carrier is a sustained release mixture.

Composition comprising a metal oxide and a metal oxide

According to another aspect, the invention provides a site-specific, sustained-release pharmaceutical composition for treating hematoma expansion or recurrent rebleeding resulting from a hemorrhagic condition in the brain, the pharmaceutical composition comprising: (a) a therapeutically effective amount of an anti-fibrinolytic agent and (b) a pharmaceutically acceptable carrier, wherein the pharmaceutically acceptable carrier is a sustained release carrier.

According to one embodiment, the hemorrhagic brain condition is rebleeding following traumatic brain injury.

According to another embodiment, the hemorrhagic brain condition is chronic SDH.

According to another embodiment, the hemorrhagic brain condition is ICH.

According to another embodiment, the intracerebral hematoma is idiopathic ICH.

According to another embodiment, the intracerebral hematoma is traumatic ICH.

According to another embodiment, the hemorrhagic brain condition is rebleeding after a craniotomy.

According to another embodiment, craniotomy procedures are performed to treat brain cancer.

According to another embodiment, a craniotomy procedure is performed to treat an intracerebral vascular abnormality.

According to another embodiment, a craniotomy procedure is performed to treat a cerebral aneurysm.

According to another embodiment, the anti-fibrinolytic agent is-aminocaproic acid (amistar).

According to another embodiment, the anti-fibrinolytic agent is factor VII.

According to another embodiment, the factor VII is a recombinant factor VII.

According to another embodiment, the anti-fibrinolytic agent is tranexamic acid.

According to another embodiment, the anti-fibrinolytic agent is aprotinin.

According to another embodiment, the pharmaceutically acceptable carrier is a controlled release carrier.

According to another embodiment, the pharmaceutically acceptable carrier is a sustained release carrier.

According to another embodiment, the anti-fibrinolytic agent is embedded in the sustained-release carrier.

According to another embodiment, the anti-fibrinolytic agent is coated on the sustained-release carrier.

According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration.

According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration.

According to another embodiment, the sustained release carrier comprises microparticles.

According to another embodiment, the sustained release carrier comprises nanoparticles.

According to another embodiment, the sustained release carrier comprises a biodegradable polymer.

According to another embodiment, the biodegradable polymer is a synthetic polymer.

According to another embodiment, the biodegradable polymer is a naturally occurring polymer.

According to another embodiment, the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof.

According to another embodiment, the synthetic polymer is polyglycolic acid (PGA).

According to another embodiment, the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA) or polycaprolactone.

According to another embodiment, the sustained release carrier is a hydrogel.

According to another embodiment, the naturally occurring polymer is a protein polymer.

According to another embodiment, the protein polymer is synthesized from a self-assembled protein polymer comprising fibroin, elastin, collagen, or a combination thereof.

According to another embodiment, the naturally occurring polymer is a naturally occurring polysaccharide.

According to another embodiment, the naturally occurring polymer comprises hyaluronic acid.

According to another embodiment, the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

The therapeutic agent in the composition is administered in a therapeutically effective amount. In combination with the teachings provided herein, by selecting and balancing among various active compounds a variety of factors such as potency, relative bioavailability, patient weight, severity of adverse side effects, and preferred mode of administration, prophylactic or therapeutic treatment regimens can be designed that do not cause substantial toxicity, but yet are effective in treating a particular subject. The effective amount for any particular application may vary depending on such factors as the disease or condition being treated, the particular therapeutic agent being administered, the size of the subject, or the severity of the disease or disorder. One of ordinary skill in the art can empirically determine the effective amount of a particular therapeutic agent without undue experimentation. It is generally preferred that the maximum dose, i.e., the highest safe dose according to some medical judgment, should be used. The terms "dose" and "dose" are used interchangeably herein.

For any of the compounds described herein, a therapeutically effective amount can be initially determined from preliminary in vitro studies and/or animal models. Therapeutically effective dosages of therapeutic agents that have been tested in humans and compounds known to exhibit similar pharmacological activity (e.g., other related active agents) can also be determined from human data. The dose applied may be adjusted based on the relative bioavailability and potency of the administered compound. It is well within the ability of one of ordinary skill in the art to adjust dosages to maximum efficacy based on the methods described above and other methods as are well known in the art.

Formulations of the therapeutic agents may be administered in pharmaceutically acceptable solutions which may conventionally contain pharmaceutically acceptable concentrations of salts, buffers, preservatives, compatible carriers, adjuvants and optionally other therapeutic ingredients.

According to some embodiments, the composition of the invention comprising a therapeutic agent may further comprise one or more additional compatible active ingredients.

For therapeutic use, an effective amount of a therapeutic agent can be administered to a subject by any mode of delivering the therapeutic agent to the desired surface. Administration of such pharmaceutical compositions may be accomplished by any means known to those skilled in the art. Routes of administration include, but are not limited to, subdural, intracerebral, intrathecal, intraarterial, parenteral (e.g., intravenous), or intramuscular. The therapeutic agent may be delivered to the subject during surgery to treat an underlying disease or side effect, such as chronic SDH, ICH, or during other surgery.

When it is desired to deliver the therapeutic agent locally, it can be formulated for parenteral administration by injection (e.g., by bolus injection or continuous infusion). Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Pharmaceutical formulations for parenteral administration include aqueous solutions of the active compounds in water-soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents that increase the solubility of the compounds to allow for the preparation of highly concentrated solutions. Alternatively, the active compound may be in powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.

The pharmaceutical compositions may also contain suitable solid or gel phase carriers or excipients. Examples of such carriers or excipients include, but are not limited to, calcium carbonate, calcium phosphate, various sugars, starch, cellulose derivatives, gelatin, and polymers such as polyethylene glycol.

Suitable liquid or solid pharmaceutical product forms are, for example, microencapsulated and, if desired, microencapsulated together with one or more excipients, spiral-shaped (encochleared), coated onto microscopic gold particles, contained in liposomes, as pellets for implantation in tissue or dried onto the object to be rubbed into the tissue. Such pharmaceutical compositions may also be in the form of: granules, beads, powders, tablets, coated tablets, (micro) capsules, suppositories, syrups, emulsions, suspensions, creams, drops or preparations with delayed release of the active compound in which excipients and additives and/or adjuvants, such as disintegrants, binders, coatings, swelling agents, lubricants or solubilizers, are customarily used as described above. The pharmaceutical compositions are suitable for use in a variety of drug delivery systems. For a brief review of drug delivery methods, see Langer 1990Science 249, 1527-.

The therapeutic agent may be administered in the form of a pharmaceutically acceptable salt. For use in medicine, the salts should be pharmaceutically acceptable, but non-pharmaceutically acceptable salts may conveniently be used to prepare pharmaceutically acceptable salts thereof. Such salts include, but are not limited to, those prepared from the following acids: hydrochloric, hydrobromic, sulfuric, nitric, phosphoric, maleic, acetic, salicylic, p-toluenesulfonic, tartaric, citric, methanesulfonic, formic, malonic, succinic, naphthalene-2-sulfonic, and benzenesulfonic acids. In addition, such salts may be prepared as alkali metal or alkaline earth salts, for example, sodium, potassium or calcium salts of carboxylic acid groups. Pharmaceutically acceptable salts are well known in the art. Stahl et al, in Handbook of Pharmaceutical Salts: Properties, Selection, and Use (Wiley VCH, Zurich, Switzerland:2002) describe pharmaceutically acceptable Salts in detail, for example. These salts can be prepared in situ during the final isolation and purification of the compounds described in the present invention, or separately by reacting the free base function with a suitable organic acid. Representative acid addition salts include, but are not limited to, acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, bromate, iodate, 2-hydroxyethanesulfonate (isethionate), lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate (pamoate), pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, phosphate, glutamate, bicarbonate, p-toluenesulfonate and undecanoate. In addition, the basic nitrogen-containing groups may be quaternized with such substances as lower alkyl halides, for example, methyl, ethyl, propyl and butyl chlorides, bromides and iodides; dialkyl sulfates such as dimethyl, diethyl, dibutyl and diamyl sulfates; long chain halides such as decyl, lauryl, myristyl and octadecyl chlorides, bromides and iodides; aralkyl halides such as benzyl bromide and phenethyl bromide and others. Water-soluble or oil-soluble or dispersible products are thereby obtained. Examples of acids which may be used to form pharmaceutically acceptable acid addition salts include inorganic acids such as hydrochloric, hydrobromic, sulphuric and phosphoric acids, and such organic acids as oxalic, maleic, succinic and citric acids. Base addition salts are prepared in situ during the final isolation and purification of the compounds described within the scope of the present invention by reacting the carboxylic acid-containing moiety with a suitable base, such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation, or with ammonia or an organic primary, secondary or tertiary amine. Pharmaceutically acceptable salts include, but are not limited to, cationic salts based on alkali or alkaline earth metals, such as lithium, sodium, potassium, calcium, magnesium, and aluminum salts, and the like, as well as non-toxic quaternary ammonium and amine cations, including ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, and the like. Other representative organic amines useful for forming base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, piperazine and the like. Pharmaceutically acceptable salts can also be obtained using standard procedures well known in the art, for example by reacting a strongly basic compound (such as an amine) with a suitable acid which affords a physiologically acceptable anion. Alkali metal (e.g., sodium, potassium, or lithium) or alkaline earth metal (e.g., calcium or magnesium) salts of carboxylic acids may also be produced.

The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. All methods include the step of bringing into association the therapeutic agent, or a pharmaceutically acceptable salt or solvate thereof ("active compound"), with the carrier which constitutes one or more accessory ingredients. Typically, the formulation is prepared by: the active agent is uniformly and intimately admixed with liquid carriers or finely divided solid carriers or both, and the product is then shaped into the desired formulation, as desired.

The pharmaceutical substance or a pharmaceutically acceptable ester, salt, solvate, functional derivative or prodrug thereof may be mixed with other active agents which do not impair the desired effect, or with substances which supplement the desired effect. Solutions or suspensions used for parenteral, intradermal, subcutaneous, subdural, intracerebral, intrathecal or topical application may include, for example, but are not limited to, the following components: sterile diluents such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerin, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates, and substances for adjusting tonicity such as sodium chloride or dextrose. The parenteral preparation can be enclosed in ampoules (or ampoules) made of glass or plastic, disposable syringes or multiple dose vials. Specific carriers for intravenous administration are physiological saline or Phosphate Buffered Saline (PBS).

Pharmaceutical compositions for parenteral injection comprise pharmaceutically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions and sterile powders for reconstitution into injectable solutions or dispersions. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol, and the like), suitable mixtures thereof, vegetable oils (such as olive oil), and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

These compositions may also contain adjuvants, including preservatives, wetting agents, emulsifying agents, and dispersing agents. Prevention of the action of microorganisms can be ensured by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. It may also be desirable to include isotonic agents, for example, sugars, sodium chloride, and the like. Prolonged absorption of the injectable pharmaceutical form can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin.

Suspensions, in addition to the active compounds, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar, tragacanth, and mixtures thereof.

Injectable depot forms (depot forms) can be prepared by forming microencapsulated matrices of the drug in biodegradable polymers such as, but not limited to, polyesters (polyglycolides, polylactic acids, and combinations thereof), polyester polyethylene glycol copolymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, naturally occurring biopolymers, protein polymers, collagen, and polysaccharides. Depending on the ratio of drug to polymer and the nature of the particular polymer used, the rate of drug release can be controlled. Such depot formulations may be formulated with suitable polymeric or hydrophobic materials (e.g., emulsions in acceptable oils) or ion exchange resins, or as sparingly soluble derivatives, e.g., as a sparingly soluble salt. Injectable depot formulations can also be prepared by entrapping the drug in liposomes or microemulsions which are compatible with body tissues.

Polyglycolide (PGA) is a linear aliphatic polyester developed for use in sutures. Studies have reported PGA copolymers formed with trimethylene carbonate, polylactic acid (PLA) and polycaprolactone. Some of these copolymers may be formulated as microparticles for sustained drug release.

Polyester-polyethylene glycol compounds can be synthesized; these compounds are soft and can be used for drug delivery.

Poly (amino) -derived biopolymers may include, but are not limited to, those containing lactic acid and lysine as aliphatic diamines (see, e.g., U.S. patent 5,399,665) and tyrosine-derived polycarbonates and polyacrylates. Modification of the polycarbonate can change the length of the alkyl chain of such an ester (ethyl to octyl), while modification of the polyacrylate can further include changing the length of the alkyl chain of the diacid (e.g., succinic to sebacic acid), which results in a large shift in the polymer and a large flexibility in polymer properties.

Polyanhydrides are prepared by dehydrating two diacid molecules by melt polymerization (see, e.g., U.S. patent 4,757,128). These polymers degrade due to surface attack (as opposed to polyesters that degrade due to bulk attack). The release of the drug may be controlled by the hydrophilicity of the selected monomer.

Photopolymerizable biopolymers include, but are not limited to, lactic acid/polyethylene glycol/acrylate copolymers.

The term "hydrogel" refers to a substance that produces a solid, semi-solid, pseudoplastic, or plastic structure containing the necessary aqueous components to produce a gelatinous or jelly-like substance. Hydrogels generally comprise a variety of polymers, including hydrophilic polymers, acrylic acid, acrylamide, and 2-hydroxyethyl methacrylate (HEMA).

Naturally occurring biopolymers include, but are not limited to, protein polymers, collagen, polysaccharides, and photopolymerizable compounds.

Protein polymers have been synthesized from self-assembling protein polymers such as fibroin, elastin, collagen and combinations thereof.

Naturally occurring polysaccharides include, but are not limited to, chitin and its derivatives, hyaluronic acid, dextran, and cellulose (which are generally not biodegradable in the unmodified case), and Sucrose Acetate Isobutyrate (SAIB).

Chitin consists mainly of 2-acetamide-2-deoxy-D-glucosyl groups and is present in yeast, fungi and marine invertebrates (shrimps, crabs), where it is the main component of the exoskeleton. Chitin is not water soluble and deacetylated chitin (chitosan) is only soluble in acidic solutions (such as, for example, acetic acid). Studies have reported chitin derivatives that are water soluble, very high molecular weight (greater than 2 million daltons), viscoelastic, non-toxic, biocompatible, and capable of crosslinking with peroxides, glutaraldehyde, glyoxal or other aldehydes, and carbodiimides to form a gel.

Hyaluronic Acid (HA), which consists of alternating glucuronic acid and glucosamine bonds and is present in the vitreous humor, synovial fluid, umbilical cord and rooster comb of mammals, from which it can be isolated and purified, can also be produced by fermentation processes.

Topical injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use. Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a parenterally-acceptable, non-toxic diluent or solvent, for example, as a solution in 1, 3-butanediol. Among the acceptable vehicles and solvents that may be used are water, ringer's solution, u.s.p., Phosphate Buffered Saline (PBS), and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally employed or employed as a solvent or suspending medium. For this purpose, any low-irritation fixed oil may be used, including synthetic mono-or diglycerides. In addition, fatty acids, such as oleic acid, are used in the preparation of injectables.

Formulations for parenteral (including but not limited to intracerebral, subdural, subcutaneous, intradermal, intramuscular, intravenous, intrathecal and intraarticular) administration include aqueous and non-aqueous sterile injection solutions wherein the solution may contain antioxidants, buffers, bacteriostats and solutes that render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions, which may contain suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) environment with the addition of the sterile liquid carrier, for example, saline, water for injections, immediately prior to use. Ready-to-use injection solutions and suspensions can be prepared from sterile powders, granules and tablets of the aforementioned kind.

Another method of formulating the compositions described herein involves conjugating the compounds described herein with a polymer that enhances water solubility.

Examples of suitable polymers include, but are not limited to, polyethylene glycol, poly- (d-glutamic acid), poly- (l-glutamic acid), poly- (d-aspartic acid), poly- (l-aspartic acid), and copolymers thereof. Polyglutamic acid having a molecular weight between about 5,000 and about 100,000, having a molecular weight between about 20,000 and about 80,000 may be used, and polyglutamic acid having a molecular weight between about 30,000 and about 60,000 may also be used. Such polymers may be conjugated via an ester linkage to one or more hydroxyl groups of the therapeutic agent using a protocol substantially as described in U.S. patent No.5,977,163, incorporated herein by reference.

Suitable buffers include: acetic acid and salt (1-2% w/v); citric acid and salts (1-3% w/v); boric acid and salts (0.5-2.5% w/v); and phosphoric acid and salts (0.8-2% w/v). Suitable preservatives include benzalkonium chloride (0.003-0.03% w/v); chlorobutanol (0.3-0.9% w/v); nipagin ester (0.01-0.25% w/v) and thimerosal (0.004-0.02% w/v).

The pharmaceutical compositions in said invention contain a therapeutically effective amount of at least one therapeutic agent and optionally other therapeutic agents included in a pharmaceutically acceptable carrier. The components of the pharmaceutical composition can also be mixed in such a way that there are no interactions that substantially impair the desired pharmaceutical efficiency.

The therapeutic agent may also be provided in particles, ropes, or thin layers.

According to one embodiment, the therapeutic agent may be provided in particles. The particles may contain the therapeutic agent in a core surrounded by a coating, or the therapeutic agent may be dispersed throughout the particles, or the therapeutic agent may be adsorbed within the particles. The particles may have any level of release kinetics including zero order release, first order release, second order release, delayed release, sustained release, immediate release, and the like, and any combination thereof. In addition to the therapeutic agent, the particles may include any of those materials conventionally used in the medical arts, including but not limited to erodible, non-erodible, biodegradable or non-biodegradable materials or combinations thereof. The particles may be microcapsules, nanocapsules, or in some cases larger containing the therapeutic agent in solution or in a semi-solid state. The particles may be of virtually any shape. According to some embodiments, the particles that may contain, in whole or in part, at least one therapeutic agent are microparticles. According to some embodiments, the particles that may contain, in whole or in part, at least one therapeutic agent are nanoparticles.

According to another embodiment, the therapeutic agent may be provided in the cord. The cord may contain the therapeutic agent in a core surrounded by a coating, or the therapeutic agent may be dispersed throughout the cord, or the therapeutic agent may be adsorbed within the cord. Such a tether may have any level of release kinetics, including zero order release, first order release, second order release, delayed release, sustained release, immediate release, and the like, as well as any combination thereof. Such cords may include, in addition to therapeutic agents, any of those materials conventionally used in the medical arts, including but not limited to erodible, non-erodible, biodegradable, or non-biodegradable materials, or combinations thereof.

According to another embodiment, the therapeutic agent may be provided in a thin layer. The sheet may contain the therapeutic agent in the core surrounded by the coating, or the therapeutic agent may be dispersed throughout the sheet, or the therapeutic agent may be adsorbed within the sheet. Such a thin layer may have any level of release kinetics including zero order release, first order release, second order release, delayed release, sustained release, immediate release, and the like, as well as any combination thereof. In addition to the therapeutic agent, such a thin layer may also include any of those materials conventionally used in the medical arts, including but not limited to erodible, non-erodible, biodegradable, or non-biodegradable materials, or combinations thereof.

Both non-biodegradable and biodegradable polymeric materials may be used in the manufacture of particles for delivery of therapeutic agents. Such polymers may be natural or synthetic polymers. The polymer is selected based on the range of time periods required for release. Bioadhesive polymers of particular interest include bioerodible hydrogels as described by Sawhney et al in Macromolecules (1993)26, 581-587, the teachings of which are incorporated herein. These include polyesters (polyglycolide, polylactic acid, and combinations thereof), polyester polyethylene glycol copolymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, naturally occurring biopolymers, protein polymers, collagen, polysaccharides, photopolymerizable compounds, polyhyaluronic acid, casein, gelatin, gelatins, polyanhydrides, polyacrylic acids, alginates, chitosan, poly (methyl methacrylate), poly (ethyl methacrylate), poly (butyl methacrylate), poly (isobutyl methacrylate), poly (hexyl methacrylate), poly (isodecyl methacrylate), poly (lauryl methacrylate), poly (phenyl methacrylate), poly (methyl acrylate), poly (isopropyl acrylate), poly (isobutyl acrylate), and poly (octadecyl acrylate). In some embodiments, the bioadhesive polymers of the invention comprise hyaluronic acid. According to some such embodiments, the bioadhesive polymer comprises less than about 2.3% hyaluronic acid.

The therapeutic agent may be contained in a controlled release system. To prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subdural, intracerebral, subcutaneous, intrathecal or intramuscular injection. This can be achieved by using a liquid suspension of crystalline or amorphous material that is poorly water soluble. The rate of absorption of the drug is therefore dependent on its rate of dissolution, which in turn is dependent on crystal size and crystalline form. For example, according to some embodiments, the SABER comprising a high viscosity matrix component such as Sucrose Acetate Isobutyrate (SAIB)^TMThe delivery system is used to provide controlled release of the drug. (see U.S. Pat. No.5,747,058 and U.S. Pat. No.5,968,542, incorporated herein by reference). When the high viscosity SAIB is formulated with drugs, biocompatible excipients, and other additives, the resulting formulation is sufficient for easy injection using standard syringes and needlesThe liquid of (2). In-injection SABER^TMAfter formulation, the excipient diffused away, leaving a viscous reservoir.

As used herein, the term "controlled release" is intended to refer to any formulation containing a drug wherein the manner and profile of release of the drug from the formulation is controlled. This refers to immediate release formulations as well as non-immediate release formulations, and non-immediate release formulations include, but are not limited to, sustained release formulations and delayed release formulations. The term "sustained release" (also referred to as "extended release") is used herein in its conventional sense to refer to a pharmaceutical formulation that provides gradual release of the drug over an extended period of time and preferably, although not necessarily, produces substantially constant blood levels of the drug over an extended period of time. Alternatively, delayed absorption of a parenterally administered drug form is achieved by dissolving or suspending the drug in an oil vehicle. The term "delayed release" is used herein in its conventional sense to refer to a pharmaceutical formulation wherein there is a time delay between administration of the formulation and release of the drug therefrom. "delayed release" may or may not include gradual release of the drug over an extended period of time, and thus may or may not be "sustained release".

The use of long-term sustained release implants may be particularly suitable for treating chronic conditions. Long-term sustained release implants are well known to those of ordinary skill in the art and include some of the delivery systems described above.

Delivery system

According to another aspect, the invention provides a semi-solid delivery system for a therapeutic agent and a combined semi-solid, multiparticulate, therapeutic delivery system for a therapeutic agent. For example, the invention provides a delivery system that utilizes a semi-solid, biodegradable, biocompatible delivery system that is injected, deposited, or implanted in or on the body to facilitate a localized therapeutic effect. Alternatively, the invention provides biodegradable, biocompatible multiparticulates that are dispersed and suspended in a semi-solid, biodegradable, biocompatible biodegradable delivery system that is injected, deposited or implanted in or on the body to promote a local therapeutic effect.

Additionally, such a semi-solid delivery system comprises, at least in part, a biocompatible, biodegradable viscous semi-solid, wherein the semi-solid comprises incorporation and retention of a significant amount H₂O, which will eventually reach equilibrium levels in the presence of an aqueous environment. According to one embodiment, glyceryl monooleate (hereinafter GMO) is the intended semi-solid delivery system or hydrogel. However, many hydrogels, polymers, hydrocarbon compositions and fatty acid derivatives with similar physical/chemical properties in terms of viscosity/stiffness can act as semi-solid delivery systems.

According to one embodiment, this gel system is produced by heating the GMO above its melting point (40 ℃ to 50 ℃) and by adding a warm water-based buffer or electrolyte solution, such as phosphate buffer or physiological saline (which thus produces a three-dimensional structure). The aqueous-based buffer may consist of other aqueous solutions or combinations containing semi-polar solvents.

GMO provides a predominantly lipid-based hydrogel with the ability to incorporate lipophilic materials. GMO further provides an internal aqueous pathway for incorporation and delivery of hydrophilic compounds. It is recognized that at room temperature (about 25 ℃), such gel systems may exhibit different phases that include a wide range of viscosity magnitudes.

According to one embodiment, two gel system phases are used due to their characteristics at room and physiological temperatures (about 37 ℃) and pH (about 7.4). Within the two gel system phases, the first phase is of about 5% to about 15% H₂A lamellar phase having an O content and a GMO content of from about 95% to about 85%. The lamellar phase is a moderately viscous fluid that can be handled, poured and injected easily. The second phase is from about 15% to about 40% H₂A cubic phase consisting of an O content and a GMO content of about 85% to 60%. It has an equilibrium moisture content of about 35% to about 40% by weight. The term "equilibrium moisture content" as used herein refers to the maximum moisture content in the presence of excess water. The cubic phase thus incorporates from about 35 to about 40% by weight of water. The cubic phase is highly viscous. Viscosity can be measured, for example, by means of a Brookfield viscometer. Viscosity in excess of 1.2 million centipoise (cp); where 1.2 million cp is the maximum viscosity value obtainable by a cup and hammer of a Brookfield viscometer. According to some such embodiments, the therapeutic agent may be incorporated into a semi-solid, thereby providing a system for sustained continuous delivery of the drug. According to some such embodiments, the therapeutic agent comprises tranexamic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises aminocaproic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises factor VII. According to some such embodiments, the therapeutic agent comprises recombinant factor VII. According to some such embodiments, the therapeutic agent comprises aprotinin. According to some such embodiments, other therapeutic agents, biologically active agents, drugs, and inactive agents may be incorporated into the semi-solid to provide a local biological, physiological, or therapeutic effect at various release rates within the body.

According to some embodiments, an alternative semi-solid, modified formulation and production method is used, thereby altering the lipophilic nature of the semi-solid, or alternatively, the aqueous channels contained within the semi-solid. Thus, varying concentrations of multiple therapeutic agents can diffuse out of such a semi-solid at different rates, or be released therefrom over time via the aqueous channels of the semi-solid. Hydrophilic substances may be used to alter the semi-solid consistency or therapeutic agent release by altering the viscosity, fluidity, surface tension or polarity of the aqueous component. For example, when heated and an aqueous component is added, Glycerol Monostearate (GMS), which is structurally identical to GMO except for double bonds instead of single bonds at the 9 th and 10 th carbons of the fatty acid moiety, does not form a gel as does GMO. However, because GMS is a surfactant, GMS is in H₂Miscible in O up to about 20% w/w. The term "surfactant" as used herein means therefore at H₂In O toA limited concentration of miscible surfactant, and a polar material. When heated and stirred, 80%>H₂The O/20% GMS combination produced spreadable pastes with similar consistency to hand sanitizer. This paste was then combined with molten GMO to form a cubic phase gel with a high viscosity as described previously. According to some such embodiments, the therapeutic agent comprises tranexamic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises aminocaproic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises factor VII or recombinant factor VII. According to some such embodiments, the therapeutic agent comprises aprotinin.

According to another embodiment, the hydrolyzed gelatin is a commercially available Gelfoam^TMFor modifying the aqueous composition. Gelfoam in a concentration of about 6.25% to 12.50% by weight^TMCan be placed in a concentration of about 93.75% to 87.50% by weight of H, respectively₂O or other water-based buffer. While heating and stirring, H₂O (or other aqueous buffer)/Gelfoam^TMThe combination produced a viscous gel-like mass. The resulting material combines with GMO, so the product thus formed swells and forms a highly viscous translucent gel of less ductility compared to neat GMO gel alone.

According to another embodiment, polyethylene glycol (PEG) may be used to modify the aqueous component to aid in drug solubilization. PEG concentrations of about 0.5% to 40% by weight (depending on the PEG molecular weight) are placed in H concentrations of about 99.5% to 60% by weight, respectively₂O or other water-based buffer. While heating and stirring, H₂The O (or other aqueous buffer)/PEG combination yields viscous liquids to semi-solid materials. The resulting material was combined with GMO, so the product thus formed swelled and a highly viscous gel was formed.

Without being limited by theory, for example, the therapeutic agent is released from the semi-solid via diffusion, conceivably in a biphasic manner. The first phase includes, for example, lipophilic drugs contained within the lipophilic membrane from which diffusion into the aqueous channels occurs. The second phase includes diffusion of the drug from the aqueous channel into the external environment. Having lipophilic properties, this drug can self-orient itself within the proposed lipid bilayer structure within the GMO inner gel segment. Thus, incorporation of more than about 7.5% by weight of the drug into the GMO results in loss of integrity of this three-dimensional structure, so that the gel system no longer maintains the semi-solid cubic phase and reverses to a viscous lamellar phase liquid. According to some such embodiments, the therapeutic agent comprises tranexamic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises aminocaproic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises factor VII or recombinant factor VII. According to some such embodiments, the therapeutic agent comprises aprotinin. According to another embodiment, about 1 to about 45% by weight of the therapeutic agent is incorporated into the GMO gel at physiological temperatures without disrupting the normal three-dimensional structure. Thus, this system has the ability to produce significantly increased drug dosage flexibility. Because this delivery system is malleable, it can be delivered and manipulated at the site of implantation, for example adjacent to or in chronic SDH, to adhere to and conform to the contours of walls, cavities or other voids within the body and to completely fill all voids present. Such a delivery system ensures drug distribution and uniform drug delivery throughout the implantation site. The convenience of delivering and manipulating the delivery system inside the cavity (e.g., without limitation, the surface of the brain) is facilitated by the semi-solid delivery device. The semi-solid delivery device facilitates directed and controlled delivery of the delivery system.

According to one embodiment, the multiparticulate component consists of a biocompatible, biodegradable, polymeric or non-polymeric system used to create a solid structure including, but not limited to, nonpareil, pellets, ropes, lamellae, crystals, aggregates, microparticles or nanoparticles.

According to another embodiment, the multiparticulate component comprises poly (lactic-co-glycolide) (PLGA). PLGA is a biodegradable polymeric material for controlled and prolonged therapeutic agent delivery in the body. Such delivery systems provide enhanced therapeutic efficacy and reduced overall toxicity compared to frequent regular systemic administration. Without being limited by theory, for example, PLGA systems composed of different molar ratios of monomer subunits would facilitate greater flexibility in engineering precise release profiles to modulate targeted therapeutic agent delivery by varying polymer degradation rates. According to one embodiment, the PLGA composition is sufficiently pure to be biocompatible and remain biocompatible when biodegraded. According to one embodiment, the PLGA polymer is designed and configured as microparticles having the therapeutic agent or drug embedded therein, so that the therapeutic agent is subsequently released therefrom by methods described in more detail below. According to some such embodiments, the therapeutic agent comprises aminocaproic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises tranexamic acid or an ester, salt, hydrate, solvate, functional derivative or prodrug thereof. According to some such embodiments, the therapeutic agent comprises factor VII or recombinant factor VII. According to some such embodiments, the therapeutic agent comprises aprotinin.

According to another embodiment, the multiparticulate component consists of poly d, l (lactic acid-co-caprolactone). This provides a biodegradable polymeric material for controlled and prolonged therapeutic agent delivery in the body by means of a similar drug release mechanism as PLGA polymers. According to one embodiment, also biodegradable and/or biocompatible non-polymeric materials such as GMS are used to produce multiparticulate microparticles.

According to another embodiment, the multiparticulate component is further modified by a process for encapsulating or coating the multiparticulate component with polymers of the same composition along with the same or different drug substances, different polymers along with the same or different drug substances, or by means of a multiple layering process without drug, with the same drug, with different drugs, or with multiple drug substances. This allows for the creation of a multi-layered (encapsulated) multi-particulate system with a wide variety of drug release profiles for a single or multiple agents simultaneously. According to another embodiment, a coating material that controls the rate of physical diffusion of the drug from the multiparticulates may be used alone or in combination with the preferred and contemplated embodiments described above.

According to another embodiment, the present invention provides a delivery system utilizing PLGA. Such PLGA polymers contain ester linkages which are susceptible to hydrolysis. At H₂When O permeates this PLGA polymer, the ester bond is hydrolyzed and the water soluble monomer is removed from the PLGA polymer, thus facilitating the physical release of the entrapped drug over time. According to some such embodiments, other classes of synthetic biodegradable, biocompatible polymers may be used for controlled and prolonged therapeutic agent delivery in the body, including polyanhydrides, poly (phosphate esters), polydioxanones, cellulosics, and acrylics, extended as non-limiting examples. According to some such embodiments, the non-polymeric material may be used for controlled and prolonged therapeutic agent delivery in the body, including but not limited to sterols, sucrose fatty acid esters, fatty acids, and cholesterol esters, as extended by non-limiting examples.

According to another embodiment, the invention provides a semi-solid delivery system that acts as a vehicle for topical delivery of therapeutic agents, comprising a lipophilic, hydrophilic or amphiphilic, solid or semi-solid substance, heated above its melting point and subsequently comprising a warm aqueous component, thereby producing a gelatinous composition of variable viscosity based on water content. The therapeutic agent is incorporated and dispersed into the molten lipophilic component or aqueous buffer component prior to mixing and forming the semi-solid system. The gelatinous composition is placed inside a semi-solid delivery device for subsequent placement or deposition. Because of its malleability, it can be easily delivered and manipulated by a semi-solid delivery device in the implantation site, adhering to and conforming to the contours of the implantation site, cavity or other void within the body and completely filling all voids present. Alternatively, a multiparticulate component consisting of a biocompatible polymeric or non-polymeric system is used to produce microparticles having the therapeutic agent embedded therein. Following the final processing method, the microparticles are incorporated into a semi-solid system and then placed inside a semi-solid delivery device so as to be easily implanted from the site of delivery or comparable lacunae, from which the therapeutic agent is then released by a drug release mechanism.

According to another embodiment, the SABER comprises a high viscosity matrix component such as Sucrose Acetate Isobutyrate (SAIB)^TMThe delivery system is used to provide controlled release of the drug.

Methods for treating hemorrhagic conditions of the brain

According to another aspect, the invention provides a method for treating hematoma expansion or recurrent rebleeding due to a brain hemorrhagic condition in a mammal, the method comprising the steps of:

(a) providing a pharmaceutical composition comprising:

(i) a therapeutically effective amount of an anti-fibrinolytic agent; and

(ii) a pharmaceutically acceptable carrier;

(b) administering the pharmaceutical composition of (a) into or at a distance proximal to an intracerebral hematoma; and

(c) Improving the medication result of patients.

According to one embodiment, the hemorrhagic condition results from Traumatic Brain Injury (TBI).

According to another embodiment, the hemorrhagic condition is post-surgical removal of a hematoma followed by hemorrhage.

According to another embodiment, the hemorrhagic condition is a chronic subdural hematoma (SDH).

According to another embodiment, the hemorrhagic condition is an intracerebral hematoma (ICH).

According to another embodiment, the intracerebral hematoma is a spontaneous intracerebral hematoma (ICH).

According to another embodiment, the intracerebral hematoma is a traumatic intracerebral hematoma (ICH).

According to another embodiment, the hemorrhagic condition is rebleeding after craniotomy.

According to another embodiment, craniotomy procedures are performed to treat brain cancer.

According to another embodiment, a craniotomy procedure is performed to treat an intracerebral vascular abnormality.

According to another embodiment, a craniotomy procedure is performed to treat a cerebral aneurysm.

According to another embodiment, the administration is implantation.

According to another embodiment, the anti-fibrinolytic agent is-aminocaproic acid (amistar).

According to another embodiment, the anti-fibrinolytic agent is factor VII.

According to another embodiment, the factor VII is a recombinant factor VII.

According to another embodiment, the anti-fibrinolytic agent is tranexamic acid.

According to another embodiment, the anti-fibrinolytic agent is aprotinin.

According to another embodiment, the pharmaceutically acceptable carrier is a controlled release carrier.

According to another embodiment, the pharmaceutically acceptable carrier is a sustained release carrier.

According to another embodiment, the anti-fibrinolytic agent is embedded in the sustained-release carrier.

According to another embodiment, the anti-fibrinolytic agent is coated on the sustained-release carrier.

According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration.

According to another embodiment, the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration.

According to another embodiment, the sustained release carrier is a microparticle.

According to another embodiment, the sustained release carrier is a nanoparticle.

According to another embodiment, the sustained release carrier comprises a biodegradable polymer.

According to another embodiment, the biodegradable polymer is a synthetic polymer.

According to another embodiment, the biodegradable polymer is a naturally occurring polymer.

According to another embodiment, the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof.

According to another embodiment, the synthetic polymer is polyglycolic acid (PGA).

According to another embodiment, the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA) or polycaprolactone.

According to another embodiment, the sustained release carrier is a hydrogel.

According to another embodiment, the naturally occurring biopolymer is a protein polymer.

According to another embodiment, the naturally occurring polymer comprises hyaluronic acid.

According to another embodiment, the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 9 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 8 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 7 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 6 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 5 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 4 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 3 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 2 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.5mm to about 1 mm.

According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 9 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 8 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 7 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 6 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 5 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 4 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 3 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 2 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.6mm to about 1 mm.

According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 9 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 8 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 7 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 6 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 5 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 4 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 3 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 2 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.7mm to about 1 mm.

According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 9 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 8 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 7 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 6 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 5 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 4 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 3 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 2 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.8mm to about 1 mm.

According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 9 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 8 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 7 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 6 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 5 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 4 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 3 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 2 mm. According to another embodiment, the distance proximal to the hematoma is from about 0.9mm to about 1 mm.

According to another embodiment, the distance proximal to the hematoma is from about 1mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 2mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 3mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 4mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 5mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 5mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 6mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 7mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 8mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9.5mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9.6mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9.7mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9.8mm to about 10 mm. According to another embodiment, the distance proximal to the hematoma is from about 9.9mm to about 10 mm.

According to another embodiment, the pharmaceutical composition exhibits a localized pharmacological effect.

According to another embodiment, the pharmaceutical composition exhibits a pharmacological effect through the brain.

According to another embodiment, the therapeutically effective amount is from about 0.000001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000003mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.0003mg to about 10g per kg of body weight. According to another embodiment, the therapeutically effective amount is from about 0.00004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.0001mg/kg body weight to about 10g/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.0005mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.001mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.005mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.01mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.1mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 1mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 10mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 20mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 30mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 40mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 50mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 60mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 70mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 80mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 90mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 100mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 110mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 120mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 130mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 140mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 150mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 160mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 170mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 180mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 190mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 200mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 250mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 500mg/kg body weight.

V methods for treating the severity of rebleeding after surgical clearance of hematoma due to hemorrhagic encephalopathy

Additionally, the invention provides a method of preventing or reducing the severity of rebleeding after surgical clearance of a hematoma resulting from a hemorrhagic brain disease, the method comprising the step of (a) implanting a pharmaceutical composition comprising (i) a therapeutically effective amount of an active agent and (ii) a coating at a distance proximate to at least one edge of the hematoma, wherein the active agent produces a localized pharmacological effect. According to one embodiment, the hemorrhagic brain condition is chronic SDH. According to another embodiment, the hemorrhagic brain condition is an intracerebral hematoma.

According to one embodiment, the hemorrhagic condition of the brain is chronic SDH.

According to another embodiment, the hemorrhagic condition of the brain is intracranial hemorrhage.

According to another embodiment, the condition of the brain is a cavity. According to some such embodiments, the cavity is a cavity created after removal of a tumor. According to some such embodiments, the cavity is a cavity created after removal of the infection. According to some such embodiments, the cavity is a cavity created after removal of a portion of the brain. According to some such embodiments, the cavity is a cavity created after removal of a vascular malformation of the brain.

According to another embodiment, the active agent comprises aminocaproic acid. According to another embodiment, the active agent comprises tranexamic acid. According to another embodiment, the active agent comprises factor VII. According to another embodiment, the active agent comprises recombinant factor VII. According to another embodiment, the active agent comprises aprotinin. According to another embodiment, the active agent comprises antiplasmin. According to another embodiment, the active agent comprises fibrin fragment D. According to another embodiment, the active agent comprises vitamin K. According to another embodiment, the active agent comprises vitamin K1. According to another embodiment, the active agent comprises vitamin K2. According to another embodiment, the active agent comprises vitamin K3. According to another embodiment, the active agent comprises 4-aminomethylbenzoic acid or an ester, salt, hydrate, solvate, prodrug or functional derivative thereof.

According to one embodiment, the pharmaceutical composition comprises a gel. According to another embodiment, the pharmaceutical composition comprises a slow release solid. According to another embodiment, the pharmaceutical composition comprises a semi-solid compound.

According to another embodiment, the pharmaceutical composition is a controlled release pharmaceutical composition. According to another embodiment, the pharmaceutical composition is a sustained release pharmaceutical composition. According to another embodiment, the pharmaceutical composition is a sustained release pharmaceutical composition.

According to another embodiment, the coating comprises at least one pharmaceutically acceptable polymer. According to some embodiments, the coating forms a matrix with the active agent, wherein the active agent has a desired release profile. According to some embodiments, the coating is mixed with the active agent during the granulation phase of the formulation.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 9mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 8mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 7mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 6mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 5mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 4mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 3mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 2mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.5mm to about 1mm from the at least one edge of the hematoma.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 9mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 8mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 7mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 6mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 5mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 4mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 3mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 2mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.6mm to about 1mm from the at least one edge of the hematoma.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 9mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 8mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 7mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 6mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 5mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 4mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 3mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 2mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.7mm to about 1mm from the at least one edge of the hematoma.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 9mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 8mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 7mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 6mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 5mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 4mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 3mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 2mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.8mm to about 1mm from the at least one edge of the hematoma.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 9mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 8mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 7mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 6mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 5mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 4mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 3mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 2mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 0.9mm to about 1mm from the at least one edge of the hematoma.

According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 1mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 2mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 3mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 4mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 5mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 6mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 7mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 8mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9.5mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9.6mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9.7mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9.8mm to about 10mm from the at least one edge of the hematoma. According to another embodiment, the distance proximate to the at least one edge of the hematoma is from about 9.9mm to about 10mm from the at least one edge of the hematoma.

According to another embodiment, the pharmaceutical composition is implanted by surgical injection.

According to another embodiment, a therapeutically effective amount of the composition is from about 0.000001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000003mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.000009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00001mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00002mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.0003mg to about 10g per kg of body weight. According to another embodiment, the therapeutically effective amount is from about 0.00004mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00005mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00006mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00007mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00008mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.00009mg/kg body weight to about 10g/kg body weight. According to another embodiment, the therapeutically effective amount is from about 0.0001mg/kg body weight to about 10g/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.0005mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.001mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.005mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.01mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 0.1mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 1mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 10mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 20mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 30mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 40mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 50mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 60mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 70mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 80mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 90mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 100mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 110mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 120mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 130mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 140mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 150mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 160mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 170mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 180mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 190mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 200mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 250mg/kg body weight. According to some such embodiments, the therapeutically effective amount is about 500mg/kg body weight.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited.

Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range is included in the invention, or any other stated or intervening value in that stated range. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges is also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the stated limits, ranges excluding any two of those included limits are also included in the invention.

It must be noted that, as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. All technical and scientific terms used herein have the same meaning.

The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently verified.

The described invention may be embodied in other specific forms without departing from the spirit or essential characteristics thereof, and, accordingly, reference should be made to the appended claims, rather than to the foregoing specification, as indicated the scope of the invention.

Examples

The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the present invention, and are not intended to limit the scope of what the inventors regard as their invention nor are they intended to suggest that the experiments below are all or the only experiments performed. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperature, etc.) but some experimental errors and deviations should be accounted for. Unless otherwise indicated, parts are parts by weight, molecular weight is weight average molecular weight, temperature is in degrees celsius, and pressure is at or near atmospheric.

Example 1 non-human animal model of Chronic subdural hematoma (SDH)

Meninges of spinal cord

The dura mater (dura mater spinalis; spinal dural dura) that forms a loose sheath around the spinal cord (medulla spinalis) represents only the inner or meningeal layer of the dura mater; the outer layer or endosteal layer is stopped at the position of the large hole of the pillow, and the position of the large hole of the pillow is occupied by the periosteum which is lined with the vertebral canal. The dura mater is separated from the arachnoid membrane by a potential cavity ("subdural space"); the two membranes are in fact in contact with each other except where they are separated by a minute amount of fluid that acts to wet the side-by-side surfaces. The dura mater is separated from the walls of the spinal canal by a compartment ("epidural space") containing a volume of loose cellular tissue and venous plexus; the location of these veins between the dura mater and the periosteum of the vertebrae thus corresponds to the location of the cranial sinuses between the meningeal layer and the endosteal layer of the dura mater. By means of the sleeve-like sleeve, the dura mater is attached to the circumference of the occipital foramen, to the second and third cervical vertebrae, and to the posterior longitudinal ligament, in particular near the lower end of the spinal canal. The subdural space terminates at the inferior border of the second sacral vertebra; below this level, the dura mater tightly surrounds the terminal filament (the elongated thread-like extension of the spinal cord (the rearmost part of the pia mater) below the lumbar nerve origin) and descends to the coccygeal back, where it mixes with the periosteum. The sheath of the dura mater is much larger than necessary to contain its contents, and its size is larger in the neck and lumbar regions than in the thorax. Two openings can be seen on each side through the two roots of the respective spinal nerve, while the dura continues over these nerves in the form of tubular extensions as the nerves pass through the intervertebral foramen. These extensions are short in the upper half of the spine but become progressively longer in the inferior direction, forming numerous fibromembranous canals that surround the lower spinal nerves and are contained within the spinal canal.

The dura mater is similar in structure to the meningeal or supporting layer of the dura mater. It consists of white fibrous and elastic tissues arranged in bundles or layers, which for the most part are parallel to each other and have a longitudinal arrangement. The inner surface of which is smooth and covered by a mesothelial layer. It is rarely supplied with blood vessels and a few nerves have been traced back into it.

The spinal part of the arachnoid membrane (the spinal arachnoid membrane) is a thin, fragile tubular membrane that loosely surrounds the spinal cord. Above, it engages the arachnoid brain; below, it widens and surrounds the cauda equina (from the spinal nerve root bundle that passes through the lumbar and sacral spinal nerves in the inferior subarachnoid space in the first lumbar inferior vertebral canal) and nerves that proceed from it. It is separated from the dura mater by the subdural space, but the lacuna is intermittently traversed by isolated connective tissue trabeculae, which are the largest in number on the posterior surface of the spinal cord.

The spinal cord portion of the subarachnoid space is a wide compartment and is largest in the inferior half of the vertebral canal, where the arachnoid space surrounds the nerve forming the cauda equina. Above, it is joined with the subarachnoid space of the cranium; below, it terminates at the level of the inferior border of the second sacral vertebra. It is partially divided by the mediastinum (subarachnoid septum), which connects the arachnoid to the pia mater opposite the posterior median sulcus of the spinal cord and forms a partition, incomplete and perforated above, but more intact in the thoracic region. The subarachnoid space is further subdivided by dentate ligaments (serrated shelf-like extensions of the pia mater protruding from either side of the cervical and thoracic spinal cords at the frontal plane).

The pia of the dura mater (pia mater), which is thicker, stronger and less vascular than the pia mater, consists of two layers. The outer or additional soft membrane layer consists of a majority of longitudinally aligned bundles of connective tissue fibers. Between these layers are the fissured lacunae, which is in communication with the subarachnoid space, and numerous blood vessels, which are surrounded in the perivascular lymphatic sheath. The pia mater covers the entire surface of the spinal cord and adheres very closely thereto; at the front, the pia mater sends a protrusion that folds back into the anterior fissure. A longitudinal fiber band (anterior spinal fiber cord) extending along the midline of the anterior surface; with the dentate ligaments on either side. Below the spinal cord cone (the end of the spinal cord), the pia mater continues as a thin, long filament (the terminal filament) that descends through the center of the nerve block, forming the cauda equina. It blends with the level of the dura mater at the inferior border of the second sacral vertebra and extends down as far as the base of the coccyx where it fuses with the periosteum. The pia mater assists in maintaining the spinal cord in place during trunk movements, and in this case is referred to as the spinal central ligament.

Experimental operating scheme

Chronic SDH will be formed in a mouse model. Adult C57BL6, CD1, or other suitable mouse strains weighing 25-35 grams will be sedated intra-abdominally with ketamine (100mg/kg) and xylazine (10 mg/kg). Body temperature will be maintained at 37 ℃ with a rectal temperature probe and a thermostatic pad. Donor mice (or rats) will be subjected to intra-abdominal anesthesia (ketamine (100mg/kg) and xylazine (10mg/kg), and blood was collected from the external jugular vein using a No. 21 catheter, the collected allogeneic (or xenogeneic) blood was immediately injected (in 2ml, 5ml or 10ml volumes) into the subcutaneous compartment of the thoracic vertebrae of the recipient mice, the initial hematoma would be immediately measured using a digital caliper, the hematoma measurement would be repeated, with each measurement being performed independently by a different trained technician, and subsequent measurements will be taken every two days after injection, three times a day (boundary day.) the animals are placed under general anesthesia and then perfused with NaCl (0.9%, 50ml) and paraformaldehyde (4%, 120ml Phosphate Buffered Saline (PBS) solution) for 20 minutes via the left ventricle when (i) the hematoma size begins to decrease or (ii) the hematoma size subsequently expands for approximately 6 days.

The hematoma will be dissected extensively from the thoracic vertebra so that it remains intact, and then sliced coronally through the midline. For histological studies, tissue blocks will be embedded with paraffin, sectioned (10 μm), stained with hematoxylin and eosin, and examined quantitatively (i) for granulation and tissue thickness and qualitatively (ii) for inflammatory cells, neovascularization, macrophages, eosinophils, and other histological features.

A portion of the coronal section will be immunohistochemically performed. Tissues will be fixed (4% paraformaldehyde) overnight, buried in a suitable medium (e.g., optimal cutting power (OCT) medium (biogenet, Markham, Ontario)) and frozen on dry ice. Subsequent sections (10urn) will be performed with a cryostat and sections blocked (phosphate buffered saline (PBS) containing 10% normal goat serum, 1% bovine serum albumin and 0.1% sodium azide) and permeabilized (0.3% Triton X-100) for 1 hour with gentle shaking. The first antibody used in immunofluorescence procedures (commercially available from suppliers such as Abeam, Cambridge, MA) will be antibodies against macrophage (CD68), tissue-and urokinase-type plasminogen activator, TNF α, IL-6 and IL-8. Sections were incubated with primary antibody in PBS with 1% BSA, followed by washing and application of secondary antibody (e.g., Alexa Fluor 568(Invitrogen, Carlsbad, CA) goat antibody against appropriate species and isotype). After the final wash, sections were protected with coverslips with anti-fade coverslip media sealed with nail polish and stored at 4 ℃.

The sections will be viewed on a confocal microscope with a Charge Coupled Device (CCD) camera. Consistent acquisition parameters (exposure time, laser power intensity, and pinhole size) will be used. The cell number will be quantified from randomly selected haematoma wall images (n ═ 10) using an unbiased counting rule and 2 observers counting stained cells (no knowledge of experimental groupings).

If each hematoma is ellipsoid, the hematoma volume is calculated as (volume 4/3 π ABC, where A, B and C are 3 orthogonal halvesRadius (orthogonal radii)). Comparisons between groups will be performed by analysis of variance (ANOVA) or ANOVA for repeated measures (if appropriate) followed by Turkey multiple comparison test. The comparison between the two magnitudes will be made by the intra-group paired t-test and the unpaired t-test between the different groups. Linear regression will be performed using least squares and fitting to the curve will use SigmaPlut (Statistical Package for the Social Sciences SPSS)]Chicago, IL) or Stata (Stata Corp., College Station, TX) by the Levenburg-Marquardt algorithm. Nonparametric quantities will pass x²Or Fisher's exact test.

Example 3 Another model of Chronic subdural hematoma in mice

In another model, chronic SDH can be generated by a single intraperitoneal injection of 6-aminonicotinamide (25mg/kg body weight) in neonatal mice on postnatal day 5. Some number of these mice can spontaneously develop spontaneous SDH after 20 or more days after injection. The control can be injected with the same volume of saline. Hematoma was assessed after left ventricular perfusion of anesthetized mice with 50ml 0.9% NaCl followed by 120ml 4% paraformaldehyde in Phosphate Buffered Saline (PBS) over 20 minutes (fig. 9).

Example 4 time course and fluid analysis of Chronic subdural hematomas in mice

The time course of the formation of the chronic SDH and the fluid analysis of the chronic SDH will be performed. A volume of allogeneic blood (2ml, 5ml or 10ml) will be injected into the subcutaneous compartment of the thoracic vertebrae of the recipient mouse and the time to hematoma formation will be monitored. Immediately prior to sacrifice, hematoma fluid was aspirated into a siliconized tube containing protamine sulfate and ethylenediaminetetraacetic acid (EDTA). Control venous blood will be obtained from the femoral vein. All samples will be centrifuged (3000 rpm 10 min) and the supernatant subsequently removed and stored (-80 ℃). Using commercially available ELISA kits (from, e.g., R and D Systems, Minneapolis, MN or America)an Diagnostics Stamford, CT), the sample will be analyzed α₂Antiplasmin, plasmin- α₂Antiplasmin complex, IL-6, IL-8 and TNF α.

Example 5 knock-out of mice to manipulate fibrinolysis

Wild-type mice and mice that have been knocked-out for t-PA and α 2-antiplasmin will receive injections of allogeneic blood into the subcutaneous compartment above the thoracic vertebra the hematoma volume will be measured by digital calipers at the time of euthanasia, the hematoma will be evaluated histologically and immunochemically and the hematoma fluid will be analyzed for fibrinolytic markers such as plasminogen activator, α₂Antiplasmin, plasmin- α 2-antiplasmin complex.

Example 6 pharmacological manipulation of fibrin

A pharmaceutical composition comprising PLGA or similar biodegradable polymer together with tranexamic acid will be synthesized. Briefly, 6 tranexamic acid and PLGA formulations with variable release kinetics will be synthesized. The release kinetics will be modified by changing the composition of the PLGA. The different formulations will be tested for their in vitro release kinetics in a mouse model. The release of tranexamic acid will be measured by thromboelastography.

Example 7 rat model of ICH

The preparation will be tested in the ICH rat collagenase model, which is more likely to be accompanied by persistent bleeding than other models in which a single blood injection is used (Eiger, b. et al, j.stroke cerebrovasc. dis.7:10,1998).

Groups of 10 rats per group will receive injections of collagenase along with the formulation of the invention into the caudate nucleus. Rats will be subjected to surgery under general anesthesia induced by intraperitoneal injections of ketamine (90mg/kg) and xylazine (10 mg/kg). The scalpel will be prepared aseptically with povidone-iodine and the anterior to coronal sagittal point [ the point of cranium at the junction of the sagittal and coronal sutures ] will be the median occipital incision. The skull would be exposed and a hole would be drilled above the caudate nucleus with a 1mm drill. The tail artery was cannulated with PE10 tubing sutured in place and connected to a pressure transducer with a rigid tube filled with 0.9% NaCl. Body temperature will be maintained and monitored with a heating pad and rectal temperature probe. Baseline blood pressure and body temperature will be recorded for 15 minutes, after which collagenase (type IV, 0.15 units, e.g. from Worthington Biochemicals, Lakewood, NJ) will be injected with or without controls or formulations.

Animals will be euthanized from 1 to 10 days post ICH, and brains removed and examined gross and histologically. The brain will be fixed in formalin and then hemispherical sections will be excised, embedded in paraffin, sectioned and stained with hematoxylin and eosin. The size of the hematoma will be measured by an integration method using standard morphometric methods. The sample will be examined and the extent of brain parenchymal inflammation will be assessed by the overall appearance, number of infiltrating multinucleated neutrophils in the connective tissue surrounding the injection site, number of monocytes, presence of any necrosis, capillary hyperplasia and fibrosis. Changes will be assessed on a three-component scale, where 1 ═ neutrophil infiltration is accompanied by little or no inflammatory component; 2 ═ non-suppurative inflammation; and 3 ═ cell necrosis with acute suppurative inflammation.

The serological effect of the composition of the invention will be determined by thromboelastography on days 2 and 5.

Example 8: synthesis of sustained release antifibrinolytic agent compositions

Pharmaceutical compositions comprising a sustained release carrier such as PLGA or similar biodegradable polymers can be co-synthesized with an anti-fibrinolytic agent (e.g., -aminocaproic acid (amica), factor VII (wild type or recombinant), aprotinin, tranexamic acid). For example, sustained release antifibrinolytic agents can be embedded in or coated on microparticles (50 microns) made of poly (D, L-lactide-co-glycolide).

To determine the pharmacokinetics of such sustained release anti-fibrinolytic agent compositions, 6 formulations of fibrinolytic agent and PLGA with variable release kinetics can be synthesized. The release kinetics can be modified by varying the composition of the PLGA. Subsequently, the ability of the different formulations to reduce ICH volume in an ICH rat model was tested when administered in vivo with collagenase. The release of fibrinolytic agents can be measured by thromboelastography.

Example 9: treatment of hematoma expansion or recurrent bleeding by site-specific administration of sustained release antifibrinolytic agent compositions

Patients diagnosed with intracerebral hemorrhage by CT scanning may be treated by implanting a plurality of sustained release microparticles comprising an anti-fibrinolytic agent (-aminocaproic acid, factor VII, tranexamic acid, or aprotinin). For example, multiple sustained release microparticles (50 microns) made of poly (D, L-lactide-co-glycolide) containing aminocaproic acid can be implanted within or near a hematoma in the brain of a patient within 72 hours after baseline CT scan or no later than 96 hours after onset of symptoms. The administration of each anti-fibrinolytic agent may be based on estimated body weight. Once implanted, the sustained release microparticles may release the anti-fibrinolytic agent for a particular time window (e.g., 3-5 days) when most of the rebleeding occurs.

Following implantation, follow-up CT scans were performed at target intervals of 24 hours (range 21 to 27 hours) and 72 hours (range 66 to 78 hours). The digital CT data can be analyzed by a neuroradiology specialist with analytical software (Mayo clinic). The volume of intracerebral hemorrhage, intracerebroventricular hemorrhage, and edema can be calculated using a computerized quadrature technique.

While the invention has been described with reference to specific embodiments thereof, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the true spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation, material, composition of matter, method, process step or steps, to the objective spirit and scope of the present invention. All such modifications are intended to be within the scope of the appended claims.

1. A method for treating hematoma expansion or recurrent bleeding resulting from a hemorrhagic condition in the brain, the method comprising:

(a) providing a pharmaceutical composition comprising

(i) A therapeutically effective amount of at least one anti-fibrinolytic agent, and

(ii) a pharmaceutically acceptable carrier;

(b) administering the pharmaceutical composition into or at a distance proximal to an intracerebral hematoma; and

(c) improving the medication result of patients.

2. The method according to embodiment 1, wherein the hemorrhagic condition results from Traumatic Brain Injury (TBI).

3. The method according to embodiment 1, wherein the hemorrhagic condition is rebleeding after surgical removal of a hematoma.

4. The method according to embodiment 1, wherein the hemorrhagic condition is a chronic subdural hematoma (SDH).

5. The method according to embodiment 1, wherein the hemorrhagic condition is an intracerebral hematoma (ICH).

6. The method of embodiment 5, wherein the intracerebral hematoma is a spontaneous intracerebral hematoma (ICH).

7. The method of embodiment 5, wherein the intracerebral hematoma is a traumatic intracerebral hematoma (ICH).

8. The method according to embodiment 1, wherein the hemorrhagic condition is rebleeding following a craniotomy procedure.

9. The method of embodiment 8, wherein the craniotomy procedure is performed to treat brain cancer.

10. The method of embodiment 8, wherein the craniotomy procedure is performed to treat a vascular abnormality in the brain.

11. The method of embodiment 8, wherein the craniotomy procedure is performed to treat a cerebral aneurysm.

12. The method of embodiment 1, wherein said administering is by implantation.

13. The method of embodiment 1, wherein the at least one anti-fibrinolytic agent is-aminocaproic acid (amica).

14. The method according to embodiment 1, wherein the at least one anti-fibrinolytic agent is factor VII.

15. The method according to embodiment 14, wherein said at least one factor VII is a recombinant factor VII.

16. The method according to embodiment 1, wherein the at least one anti-fibrinolytic agent is tranexamic acid.

17. The method of embodiment 1, wherein the at least one anti-fibrinolytic agent is aprotinin.

18. The method of embodiment 1, wherein the pharmaceutically acceptable carrier is a controlled release carrier.

19. The method of embodiment 1, wherein the pharmaceutically acceptable carrier is a sustained release carrier.

20. The method of embodiment 19, wherein the at least one anti-fibrinolytic agent is embedded in the sustained-release carrier.

21. The method according to embodiment 19, wherein the at least one anti-fibrinolytic agent is coated on the sustained-release carrier.

22. The method of embodiment 19, wherein the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration.

23. The method of embodiment 19, wherein the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration.

24. The method of embodiment 19, wherein the sustained release carrier is a microparticle.

25. The method of embodiment 19, wherein the sustained release carrier is a nanoparticle.

26. The method of embodiment 19, wherein the sustained release carrier comprises a biodegradable polymer.

27. The method of embodiment 26, wherein the biodegradable polymer is a synthetic polymer.

28. The method of embodiment 26, wherein the biodegradable polymer is a naturally occurring polymer.

29. The method of embodiment 27, wherein the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof.

30. The method of embodiment 27, wherein the synthetic polymer is polyglycolic acid (PGA).

31. The method of embodiment 27, wherein the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA), or polycaprolactone.

32. The method of embodiment 19, wherein the sustained release carrier is a hydrogel.

33. The method of embodiment 28, wherein the naturally occurring biopolymer is a protein polymer.

34. The method of embodiment 28, wherein the naturally occurring polymer comprises hyaluronic acid.

35. The method of embodiment 34, wherein the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

36. The method of embodiment 1, wherein the distance proximal to the hematoma is from about 0.5mm to about 10 mm.

37. The method of embodiment 1, wherein the pharmaceutical composition exhibits a localized pharmacological effect.

38. The method of embodiment 1, wherein the pharmaceutical composition exhibits its pharmacological effect throughout the brain.

39. A site-specific, sustained-release pharmaceutical composition for treating hematoma expansion or recurrent rebleeding resulting from a hemorrhagic condition in the brain, the pharmaceutical composition comprising:

(a) a therapeutically effective amount of at least one anti-fibrinolytic agent; and

(b) a pharmaceutically acceptable carrier, wherein the pharmaceutically acceptable carrier is a sustained release carrier.

40. The pharmaceutical composition according to embodiment 39, wherein the hemorrhagic brain condition is rebleeding following traumatic brain injury.

41. The pharmaceutical composition according to embodiment 39, wherein the hemorrhagic brain condition is a chronic subdural hematoma (SDH).

42. The pharmaceutical composition according to embodiment 39, wherein the hemorrhagic brain condition is an intracerebral hematoma (ICH).

43. The pharmaceutical composition according to embodiment 42, wherein the intracerebral hematoma is a spontaneous intracerebral hematoma (ICH).

44. The pharmaceutical composition according to embodiment 42, wherein the intracerebral hematoma is a traumatic intracerebral hematoma (ICH).

45. The pharmaceutical composition according to embodiment 39, wherein the hemorrhagic brain condition is rebleeding following a craniotomy procedure.

46. The pharmaceutical composition according to embodiment 45, wherein the craniotomy procedure is performed to treat brain cancer.

47. The pharmaceutical composition according to embodiment 45, wherein the craniotomy procedure is performed to treat a vascular malformation in the brain.

48. The composition of embodiment 45, wherein the craniotomy procedure is performed to treat a cerebral aneurysm.

49. The pharmaceutical composition according to embodiment 39, wherein the at least one anti-fibrinolytic agent is-aminocaproic acid (AMICAR).

50. The pharmaceutical composition according to embodiment 39, wherein the at least one anti-fibrinolytic agent is factor VII.

51. The pharmaceutical composition according to embodiment 50, wherein said factor VII is recombinant factor VII.

52. The pharmaceutical composition according to embodiment 39, wherein the at least one anti-fibrinolytic agent is tranexamic acid.

53. The pharmaceutical composition according to embodiment 39, wherein the at least one anti-fibrinolytic agent is aprotinin.

54. The pharmaceutical composition according to embodiment 39, wherein the pharmaceutically acceptable carrier is a controlled release carrier.

55. The pharmaceutical composition according to embodiment 39, wherein the pharmaceutically acceptable carrier is a sustained release carrier.

56. The pharmaceutical composition according to embodiment 55, wherein the at least one anti-fibrinolytic agent is embedded in the sustained-release carrier.

57. The pharmaceutical composition according to embodiment 55, wherein the at least one anti-fibrinolytic agent is coated on the sustained-release carrier.

58. The pharmaceutical composition of embodiment 55, wherein the sustained release carrier releases the anti-fibrinolytic agent for at least 21 days after administration.

59. The pharmaceutical composition of embodiment 55, wherein the sustained release carrier releases the anti-fibrinolytic agent for about 3 to 5 days after administration.

60. The pharmaceutical composition of embodiment 55, wherein the sustained release carrier comprises microparticles.

61. The pharmaceutical composition of embodiment 55, wherein the sustained release carrier comprises a nanoparticle.

62. The pharmaceutical composition according to embodiment 55, wherein the sustained-release carrier comprises a biodegradable polymer.

63. The pharmaceutical composition of embodiment 62, wherein the biodegradable polymer is a synthetic polymer.

64. The pharmaceutical composition of embodiment 62, wherein the biodegradable polymer is a naturally occurring polymer.

65. The pharmaceutical composition of embodiment 63, wherein the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, and combinations thereof.

66. The pharmaceutical composition according to embodiment 63, wherein the synthetic polymer is polyglycolic acid (PGA).

67. The pharmaceutical composition of embodiment 63, wherein the synthetic polymer is a polyglycolic acid copolymer formed with trimethylene carbonate, polylactic acid (PLA), or polycaprolactone.

68. The pharmaceutical composition according to embodiment 55, wherein the sustained-release carrier is a hydrogel.

69. The pharmaceutical composition of embodiment 64, wherein the naturally occurring polymer is a protein polymer.

70. The pharmaceutical composition of embodiment 69, wherein the protein polymer is synthesized from a self-assembled protein polymer.

71. The pharmaceutical composition of embodiment 64, wherein the naturally occurring polymer is a naturally occurring polysaccharide.

72. The pharmaceutical composition of embodiment 64, wherein the naturally occurring polymer is naturally occurring hyaluronic acid.

73. The pharmaceutical composition of embodiment 72, wherein the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

Claims (16)

1. Use of a site-specific pharmaceutical composition for the manufacture of a medicament for the treatment of hematoma enlargement or recurrent bleeding resulting from a hemorrhagic condition in the brain, said use comprising administering said composition by implantation in a cavity or compartment in the brain occupied by a hematoma, or in the subdural space on the surface of the brain,

the pharmaceutical composition comprises:

(a) a therapeutic amount of at least one anti-fibrinolytic agent, wherein the therapeutic amount is effective to reduce hematoma expansion or recurrent bleeding due to an intracerebral hemorrhagic condition when compared to an untreated control; and

(b) a biodegradable, pharmaceutically acceptable carrier, wherein the carrier comprises a copolymer of polyglycolic acid formed with polylactic acid, the carrier comprises a matrix, and wherein the carrier contains a plurality of microparticles;

wherein:

(i) the therapeutic amount of the antifibrinolytic agent is dispersed in each microparticle, within a core of the microparticle surrounded by a coating, adsorbed within the particle, or a combination thereof;

(ii) forming the biodegradable pharmaceutically acceptable carrier comprising the therapeutic amount of an anti-fibrinolytic agent to produce a solid structure selected from a rope or a lamina;

(iii) the fibrinolytic agent is continuously released for a plurality of days at the implantation position in the subarachnoid space, the subdural space of the chronic subdural hematoma or the left space after the hematoma, tumor or vascular malformation operation is removed; and

(iv) the biodegradable carrier breaks down over time, affecting the release of the anti-fibrinolytic agent into the cerebrospinal fluid circulation to achieve targeting of the therapeutic amount of the anti-fibrinolytic agent to the bleeding site, which is effective to achieve a locally high concentration of the anti-fibrinolytic agent and a lower concentration in the rest of the body, thereby reducing the risk of unwanted systemic side effects.

2. The use according to claim 1, wherein the bleeding disorder is caused by Traumatic Brain Injury (TBI).

3. The use according to claim 1, wherein the bleeding disorder is at least one disorder selected from: rebleeding after traumatic brain injury, chronic subdural hematoma (SDH), non-traumatic intracerebral hemorrhage (ICH), spontaneous intracerebral hematoma (ICH), traumatic intracerebral hematoma (ICH)), and rebleeding after craniotomy.

4. The use according to claim 3, wherein the craniotomy procedure is for the treatment of a brain tumor, an intracerebral vascular malformation, or a cerebral aneurysm.

5. The use of claim 1, wherein the anti-fibrinolytic agent is-aminocaproic acid (amiar), tranexamic acid, aprotinin, 4-aminomethylbenzoic acid, fibrin D, anti-plasmin, vitamin K, or a combination thereof.

6. The use according to claim 1, wherein the pharmaceutically acceptable carrier is a controlled or sustained release carrier.

7. The use of claim 6, wherein the anti-fibrinolytic agent is embedded in or coated on the sustained release carrier.

8. The use of claim 6, wherein the sustained release carrier is capable of releasing the anti-fibrinolytic agent for at least 21 days or about 3-5 days after administration.

9. The use of claim 6, wherein the sustained release carrier comprises nanoparticles.

10. The use of claim 6, wherein the slow release carrier comprises a biodegradable polymer or hydrogel.

11. The use according to claim 10, wherein the biodegradable polymer is a synthetic polymer or a naturally occurring polymer.

12. The use of claim 11, wherein the synthetic polymer is selected from the group consisting of polyesters, polyester polyethylene glycol polymers, polyamino-derived biopolymers, polyanhydrides, polyorthoesters, polyphosphazenes, Sucrose Acetate Isobutyrate (SAIB), photopolymerizable biopolymers, polyglycolic acid (PGA), copolymers of polyglycolic acid formed with trimethylene carbonate, polylactic acid (PLA), polycaprolactone, and combinations thereof.

13. The use of claim 11, wherein the naturally occurring polymer is a protein polymer.

14. The use of claim 11, wherein the naturally occurring polymer comprises hyaluronic acid.

15. The use of claim 14, wherein the naturally occurring polymer comprises less than 2.3% hyaluronic acid.

16. Use according to claim 5, wherein the vitamin K is selected from vitamin K1, vitamin K2 and vitamin K3.